EP2433926A1 - Pleuromutilin derivatives for use in the treatment of diseases mediated by microbes - Google Patents

Pleuromutilin derivatives for use in the treatment of diseases mediated by microbes Download PDF

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Publication number
EP2433926A1
EP2433926A1 EP10450143A EP10450143A EP2433926A1 EP 2433926 A1 EP2433926 A1 EP 2433926A1 EP 10450143 A EP10450143 A EP 10450143A EP 10450143 A EP10450143 A EP 10450143A EP 2433926 A1 EP2433926 A1 EP 2433926A1
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EP
European Patent Office
Prior art keywords
hydroxy
acetyl
mutilin
amino
diastereomer
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Application number
EP10450143A
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German (de)
French (fr)
Inventor
Rosemarie Mang
Klaus Thirring
Werner Heilmayer
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Nabriva Therapeutics AG
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Nabriva Therapeutics AG
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Publication date
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Priority to EP10450143A priority Critical patent/EP2433926A1/en
Priority to JP2013527421A priority patent/JP6125425B2/en
Priority to EP11751528.8A priority patent/EP2614049B1/en
Priority to PCT/AT2011/000342 priority patent/WO2012031307A1/en
Priority to US13/821,446 priority patent/US9278920B2/en
Priority to TW100129397A priority patent/TWI603950B/en
Publication of EP2433926A1 publication Critical patent/EP2433926A1/en
Withdrawn legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
    • C07C323/50—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton
    • C07C323/51—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton
    • C07C323/52—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton the carbon skeleton being acyclic and saturated
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/21—Esters, e.g. nitroglycerine, selenocyanates
    • A61K31/215—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids
    • A61K31/235—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids having an aromatic ring attached to a carboxyl group
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2601/00—Systems containing only non-condensed rings
    • C07C2601/04—Systems containing only non-condensed rings with a four-membered ring
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2601/00—Systems containing only non-condensed rings
    • C07C2601/06—Systems containing only non-condensed rings with a five-membered ring
    • C07C2601/08—Systems containing only non-condensed rings with a five-membered ring the ring being saturated
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2601/00—Systems containing only non-condensed rings
    • C07C2601/18—Systems containing only non-condensed rings with a ring being at least seven-membered
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2602/00—Systems containing two condensed rings
    • C07C2602/36—Systems containing two condensed rings the rings having more than two atoms in common
    • C07C2602/42—Systems containing two condensed rings the rings having more than two atoms in common the bicyclo ring system containing seven carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2602/00—Systems containing two condensed rings
    • C07C2602/36—Systems containing two condensed rings the rings having more than two atoms in common
    • C07C2602/44—Systems containing two condensed rings the rings having more than two atoms in common the bicyclo ring system containing eight carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C2603/00—Systems containing at least three condensed rings
    • C07C2603/93—Spiro compounds
    • C07C2603/95—Spiro compounds containing "not free" spiro atoms
    • C07C2603/98—Spiro compounds containing "not free" spiro atoms containing at least one ring with more than six ring members
    • C07C2603/99—Spiro compounds containing "not free" spiro atoms containing at least one ring with more than six ring members containing eight-membered rings

Definitions

  • the present invention relates to organic compounds, namely pleuromutilins.
  • Pleuromutilin a compound of formula is a naturally occurring antibiotic, e.g. produced by the basidomycetes Pleurotus mutilus and P. pasckerianus, see e.g. The Merck Index, 13th edition, item 7617 .
  • pleuromutilins having the principle ring structure of pleuromutilin and being substituted at the primary hydroxy group have been developed, e.g. as antimicrobials.
  • pleuromutilin derivatives e.g. 14-O-[(amino-hydroxy-cyclohexylsulfanyl)-acetyl]-mutilins
  • WO 2010/025482 e.g. N-acylated 14-O-[(amino-hydroxy-cyclohexylsulfanyl)-acetyl]-mutilins, are known.
  • the present invention provides compounds selected from the group of N-unsubstituted, or N-alkylated, or N-acylated 14-O-[(amino(C 0-4 )alkyl-hydroxy-cycloalkyl-and bicycloalkylsulfanyl)-acetyl]-mutilins which are
  • amino(C 0-4 )alkyl- as used herein means that the amino group is either bound to alkyl of 1 to 4 C-atoms or, in case of C 0 , directly to the (bi)cycloalkylring
  • the pesent invention provides 14-O-[(amino(C 0-4 )alkyl-hydroxy-cyclobutylsulfanyl)-acetyl]-mutilins and/or 14-O-[(amino(C 0-4 )alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins.
  • the pesent invention provides 14-O-[(amino(C 0-4 )alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins and/or 14-O-[(amino(C 0-4 )-hydroxy-cyclooctylsulfanyl)-acetyl]-mutilins.
  • the pesent invention provides 14-O-[amino(C 0-4 )alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins and/or 14-O-[amino(C 0-4 )alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins, e.g.
  • the pesent invention provides 14-O-[(amino(C 0-4 )alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins, e.g. 14-O-[(amino(C 0-4 )alkyl -hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins wherein one ring of the bicyclyic alkyl is a cyclopentyl ring, such as 14-O-[(amino(C 0-4 )alkyl-hydroxy-bicyclo[3.2.0]heptytsutfanyl)-acetyl]-mutilins and 14-O-[(amino(C 0-1 )alkyl-hydroxy-octahydropentalensulfanyl]-acetyl ⁇ -mutilins.
  • alkyl may be not present for the case of C 0 ; or may be C 1 alkyl; or may be C 2 alkyl, or may be C 3 alkyl, or may be C 4 alkyl.
  • the present invention provides a compound of formula (I) or of formula (II) wherein
  • Preferred compounds of the present invention are compounds of formula (III) or of formula (IV) wherein m, n, o, p, R 1 and R 2 are as defined above, e.g. o is 1; e.g. p is 1 or 2.
  • More preferred compounds of the present invention are compounds of formula (V) or of formula (VI) or of formula (VII) or of formula (VIII) wherein m, R 1 and R 2 are as defined as defined above.
  • the present invention provides a compound of Examples 1 to 17 below, such as a compound of formula I or II, e.g. a compound selected from the group consting of
  • a compound provided by the present invention is herein also designated as "compound(s) of (according to) the present invention".
  • a compound of the present invention includes a compound in any form, e.g. in free form, in the form of a salt, in the form of a solvate and in the form of a salt and a solvate.
  • the present invention provides a compound of the present invention in the form of a salt, e.g. and/or solvate.
  • a salt of a compound of the present invention includes preferably pharmaceutically acceptable salts, although pharmaceutically unacceptable salts are included, e.g. for preparation / isolation / purification purposes.
  • a salt of a compound of the present invention includes a base salt or an acid addition salt.
  • Pharmaceutically acceptable acid addition salts include salts of a compound of the present invention with an acid, e.g. hydrogen fumaric acid, fumaric acid, hydrogen tartaric acid, tartaric acid, hydrogen ethane-1,2-disulfonic acid, ethane-1,2-disulfonic acid, hydrogen maleic acid, maleic acid, hydrogen naphthalin-1,5-disulfonic acid, hydrogen naphthalin-1,5-disulfonic acid, acetic acid, hydrogen malic acid, malic acid, lactic acid, hydrogen succinic acid, succinic acid, salicylic acid, hydrogen azelaic acid, azelaic acid, 2-[(2,6-dichlorophenyl)-amino]-benzene acetic acid, hydrogen sulfuric acid, sulfuric acid, dihydrogen phosphoric acid, hydrogen phosphoric acid, phosphoric acid, dihydrogen citric
  • a compound of the present invention in free form may be converted into a corresponding compound in the form of a salt, and vice versa.
  • a compound of the present invention in free form or in the form of a salt and/or in the form of a solvate may be converted into a corresponding compound in free form or in the form of a salt in non-solvated form, and vice versa.
  • a compound of of the present invention may exist in the form of isomers and mixtures thereof, e.g. optical isomers, diastereoisomers, cis/trans conformers.
  • a compound of the present invention may e.g. contain asymmetric carbon atoms and may thus exist in the form of enantiomers or diastereoisomers and mixtures thereof, e.g. racemates or diastereomeric mixtures. Any asymmetric carbon atom may be present in the (R)-, (S)- or (R,S)-configuration, preferably in the (R)- or (S)-configuration.
  • the carbon atom of the cycloalkyl ring which is attached to S group, the carbon atom of the cycloalkyl ring which is attached to the OH group, and the carbon atom of the cycloalkyl ring to which the (CH 2 ) m N(R 1 R 2 ) group is attached are all asymmetric carbon atoms.
  • Substituents attached to such asymmetric carbon atoms may thus exist in (R) and (S) configuration, including mixtures thereof.
  • R 2 is substituted alkyl and that substituent is attached to a carbon atom of the side chain of such alkyl, the carbon atom to which such substituent is attached is an asymmetric carbon atom and such substituent may be in the (R)- and (S)-configuration, including mixtures thereof.
  • the configuration of asymmetric carbon atoms of the mutilin-tricyclus is preferably the same as in natural pleuromutilin.
  • Isomeric mixtures may be separated as appropriate, e.g. according, e.g. analogously, to a method as conventional, to obtain pure isomers.
  • the present invention includes a compound of the present invention in any isomeric form and in any isomeric mixture.
  • the present invention also includes tautomers of a compound of the present invention, where tautomers can exist.
  • Any compound provided according to the present invention may be obtained as appropriate, e.g. according to, e.g. analogously to a method as conventional or as described herein.
  • a compound of the present invention may be obtained by reacting a compound PLEU-Tos as defined below with an appropriate amino(C 0-4 )alkyl-hydroxy-mercapto-cyclobutane, amino(C 0-4 )alkyl-hydroxy-mercapto-cyclopentane, amino(C 0-4 )alkyl-hydroxy-mercaptocycloheptane, amino(C 0-4 )alkyl-hydroxy-mercapto-cyclooctane or amino(C 0-4 )alkyl-hydroxy-mercapto-bicycloalkane, wherein the amine group is protected, e.g. by Boc, in organic solvent, e.g.
  • polar organic solvent such as acetonitrile
  • a base and/or coupling agent such as e.g. DBN
  • deprotecting the amine group e.g. by treatment with an acid, e.g. HC1.
  • the deprotected amine group can be optionally acylated by reaction with a carboxylic acid, e.g. an amino acid, in the presence of a coupling reagent, or with activated carboxylic acid or carboxylic acid derivative, such as an anhydride of the carboxylic acid or carboxylic acid halogenide.
  • the deprotected amine group from above can be optionally alkylated, e.g. by reductive amination or via an alkylating agent.
  • amino(C 0-4 )alkyl-hydroxy-mercapto-cyclobutane, amino(C 0-4 )alkyl-hydroxy-mercaptocyclopentane, amino(C 0-4 )alkyl-hydroxy-mercapto-cycloheptane amino(C 0-4 )alkyl-hydroxy-mercapto-cyclooctane, or amino(C 0-4 )alkyl-hydroxy- mercapto-bicycloalkane,wherein the amine group is protected may be obtained from an N-protected amino(C 0-4 )alkyl cyclobutene, N-protected amino(C 0-4 )alkyl cyclopentene, N-protected amino(C 0-4 )alkyl cycloheptene, N-protected amino(C 0-4 )alkyl cyclooctene, N-protected amino(C 0-4 )alkyl bicycloalkene, respectively
  • the compounds of the present invention exhibit pharmacological activity and are therefore useful as pharmaceuticals.
  • the compounds of the present invention show antimicrobial, e.g. antibacterial, activity against gram positive bacteria, such as coagulase positive Staphylococci, e.g. Staphylococcus aureus, coagulase negative Staphylococci, e.g. Staphylococcus epidermidis, Staphylococcus haemolyticus, and Streptococci, e.g. Streptococcus pyogenes, Streptococcus pneumoniae, Enterococci, e.g. Enterococcus faecium and Listeria monocytogenes and against gram negative bacteria such as Moraxella, e.g.
  • coagulase positive Staphylococci e.g. Staphylococcus aureus
  • coagulase negative Staphylococci e.g. Staphylococcus epidermidis
  • Staphylococcus haemolyticus Staphylococc
  • Moraxella catarrhalis and Haemophilus, e.g. Haemophilus influenzae, and Legionella, e.g. Legionella pneumophila, Neisseriaceae, e.g. Neisseria gonorrhoeae, as well as against Mycoplasms, Chlamydia and obligatory anaerobes, e.g. Bacteroides fragilis, Clostridium difficile, Fusobacterium spp., and Propionibacterium spp.
  • the in vitro activity against aerobic bacteria was determined by Agar Dilution Test or Microdilution Test according to the Clinical and Laboratory Standards Institute (CLSI, former NCCLS) Document M7-A7 Vol.26, No. 2: "Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically - Approved Standard; Seventh Edition (2006 )”; and the test against anaerobic bacteria was performed according to the Clinical and Laboratory Standards Institute (CLSI, former NCCLS), Document, M11-A6, Vol. 24, No.
  • Compounds of the present invention are therefore suitable for the treatment and prevention of diseases which are mediated by microbes, e.g. by bacteria.
  • Diseases which may also be treated include e.g. diseases mediated by Helicobacter, such as Helicobacter pylori, and diseases mediated by Mycobacterium tuberculosis.
  • Diseases which may also be treated include in general inflammatory diseases, where microbes are mediating said inflammation, e.g. including acne.
  • the present invention provides a compound of the present invention for use as a pharmaceutical, preferably as an antimicrobial, such as an antibiotic, e.g. an anti-anaerobic.
  • an antimicrobial such as an antibiotic, e.g. an anti-anaerobic.
  • the present invention provides a compound of the present invention for use in acne treatment.
  • the present invention provides a compound of the present invention for use in the preparation of a medicament for the treatment of diseases, mediated by microbes, such as bacterials, for example
  • the present invention provides a method of treatment of diseases mediated by microbes which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention e.g. in the form of a pharmaceutical composition.
  • the present invention provides a method of treatment of acne which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention e.g. in the form of a pharmaceutical composition.
  • Treatment includes treatment and prophylaxis.
  • an indicated daily dosage is in the range of about 0.5 mg to 3 g of a compound of the present invention conveniently administered, for example, in divided doses up to four times a day.
  • a compound of the present invention may be administered by any conventional route, for example enterally, e.g. including nasal, buccal, rectal, oral administration; parenterally, e.g. including intravenous, intramuscular, subcutaneous administration; or topically, e.g. including epicutaneous, intranasal, intratracheal administration, e.g. in form of coated or uncoated tablets, capsules, injectable solutions or suspensions, e.g. in the form of ampoules, vials, in the form of creams, ointments, gels, pastes, inhaler powder, foams, tinctures, lipsticks, drops, sprays, or in the form of suppositories, e.g. in analogous manner to macrolides, such as erythromycins, e.g. clarithromycin or azithromycin.
  • macrolides such as erythromycins, e.g. clarithromycin or azithromycin.
  • a compound of the present invention may be administered in the form of a pharmaceutically acceptable salt, e.g. an acid addition salt or a base addition salt, e.g. a metal salt, or in free form, optionally in the form of a solvate.
  • a pharmaceutically acceptable salt e.g. an acid addition salt or a base addition salt, e.g. a metal salt
  • a compound of the present invention in the form of a salt exhibits the same order of activity as the compound in free form, optionally in the form of a solvate.
  • a compound of the present invention may be used for pharmaceutical treatment according to the present invention alone or in combination with one or more other pharmaceutically active agents.
  • Such other pharmaceutically active agents include e.g. other antibiotics and antiinflammatory agents, and, if a compound of the present invention is used in the treatment of acne, other pharmaceutically agents include furthermore agents which are active against acne.
  • Combinations include fixed combinations, in which two or more pharmaceutically active agents are in the same formulation; kits, in which two or more pharmaceutically active agents in separate formulations are sold in the same package, e.g. with instruction for coadministration; and free combinations in which the pharmaceutically active agents are packaged separately, but instruction for simultaneous or sequential administration are given.
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of the present invention in free form or in the form of a pharmaceutically acceptable salt and/or in the form of a solvate as such or in association with at least one pharmaceutical, excipient, e.g. carrier or diluent, e.g. including fillers, binders, disintegrators, flow conditioners, lubricants, sugars and sweeteners, fragrances, preservatives, stabilizers, wetting agents and/or emulsifiers, solubilizers, salts for regulating osmotic pressure and/or buffers.
  • excipient e.g. carrier or diluent
  • the present invention provides a pharmaceutical composition according to the present invention, further comprising another pharmaceutically active agent.
  • Such pharmaceutical compositions may be manufactured according, e.g. analogously, to a method as conventional, e.g. by mixing, granulating, coating, dissolving or lyophilizing processes.
  • Unit dosage form may contain, for example, from about 0.5 mg to about 2000 mg, such as 10 mg to about 500 mg.
  • the compounds of the present invention are additionally suitable as veterinary agents, e.g. veterinary active compounds, e.g. in the prophylaxis and in the treatment of microbial, e.g. bacterial diseases, in animals, such as fowl, pigs and calves, e.g., and for diluting fluids for artificial insemination and for egg-dipping techniques.
  • veterinary agents e.g. veterinary active compounds, e.g. in the prophylaxis and in the treatment of microbial, e.g. bacterial diseases, in animals, such as fowl, pigs and calves, e.g., and for diluting fluids for artificial insemination and for egg-dipping techniques.
  • the present invention provides a compound of the present invention for use as a veterinary agent.
  • the present invention provides a compound of the present invention for the preparation of a veterinary composition which is useful as a veterinary agent.
  • the present invention provides a veterinary method for the prophylaxis and the treatment of microbial, e.g. bacterial diseases which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention, e.g. in the form of a veterinary composition.
  • microbial e.g. bacterial diseases
  • mutilin refers to the IUPAC systematic name (1S, 2R, 3S, 4S, 6R, 7R, 8R, 14R)-3,6-dihydroxy-2,4,7,14-tetramethyl-4-vinyl-tricyclo[5.4.3.0 1, 8 ]tetradecan-9-one.
  • pleuromutilin derivatives are numbered in analogy to the mutilin numbering system described by H. Berner ( Berner, H.; Schulz, G.; Schneider H. Tetrahedron 1980, 36, 1807-1811 .):
  • PLEU-Tos is a compound of formula
  • PLEU is a residue of formula
  • Example 1 Step C products 780 mg was dissolved in 10 mL of DCM, 8 mL of 1 M HC1 in diethylether was added and the solution stirred for 16 h at rt. Further 2 mL of 1 M HC1 in diethylether was added and stirred for another 8 h. After evaporation of the solvent the solid residue was triturated with diethylether. After filtration, 579 mg of the Example 1 Step D products was obtained as white solid.
  • Example 2 Step B products was treated according to the method of Example 1 Step C to obtain 405 mg of Example 2 Step C products as a white solid.
  • Example 3 Step B product dissolved in 200 mL of DMF was added 27.1 g of sodium azide and the reaction mixture obtained was stirred at 60 °C for 20 h.
  • diethylether was added, the organic layer obtained was washed with water, dried and carefully evaporated using a cold water bath.
  • the crude azide product still contains DMF and is used for the next step without further purification. 18.6 g of this crude azide product was dissolved in 150 mL of THF and 5 mL of water.
  • To the mixture obtained 26.17 g of triphenylphosphine was added and the reaction mixture obtained was stirred at 80 °C for 4 h.
  • Example 3 Step C product in the form of a colorless oil.
  • Example 3 Step E products To a solution of 5.53 g of Example 3 Step E products in 80 mL of acetonitrile was added 1.56 mL of hydrazine monohydrate and the solution obtained was stirred for 30 min at rt. After addition of 25 mL of 1 M HCl the mixture obtained was stirred for 5 min, diluted with DCM and the layers were separated. The organic layer obtained was dried and solvent was evaporated. The evaporation residue obtained was dissolved in 100 mL of acetonitrile. 2.06 mL of DBN and 7.53 g of pleuromutilin tosylate was added and the solution obtained was stirred for 4 h at rt.
  • Example 3 Step F products 1g of Example 3 Step F products was dissolved in 20 mL of DCM and 5 mL of 2 M HC1 in diethylether was added. After 16 h at rt the solvent was evaporated and the evaporation residue obtained was subjected to chromatography (silica, DCM/MeOH/NH 4 OH 100:8:0.8) to obtain 155 mg of the free base
  • Example 3 Step G products in the form of a white foam.
  • Example 3 Step H 150 mg of the free base from Example 3 Step G was dissolved in 5 mL of DCM. and 1 mL of 1 M HCl in diethylether was added. After 10 min the solvent was evaporated and the hydrochloride Example 3 Step H products were obtained in the form of a white solid in a quantitative manner.
  • Example 4 Step A products in the form of a amorphous white solid.
  • Example 4 Step B products was treated according to the method of Example 3 Step G to obtain after chromatography (silica, DCM / MeOH / NH 4 OH 100:8:0.8) 906 mg of the free base Example 4 Step C products.
  • Example 4 Step C products 200 mg was dissolved in 5 mL of DCM and 1 mL of 1 M HC1 in diethylether was added. After 10 min the solvent was evaporated and 166 mg of the hydrochloride Example 4 Step D products was obtained in the form of a white amorphous solid.
  • Example 4 Step C products was dissolved in 15 mL of dry DCM containing molecular sieves. To the mixture obtained 42 mg of pivalaldehyde was added at rt and after 1 h the reaction mixture was charged with 136 mg of sodium triacetoxyborohydride and stirring was continued for another 23 h. To the mixture obtained DCM was added, the phases obtained were separated and the organic layer obtained was washed with saturated NaHCO 3 solution, dried and evaporated.
  • Example 7 Step D products in the form of a colorless oil.
  • Example 7 Step F products To a solution of 320 mg of Example 7 Step F products in 3 mL of MeOH was added 1 mL of 28% NH 4 OH at rt. After 24 h a further 1 mL of 28% NH 4 OH was added and the mixture obtained was stirred for additional 24 h. From the mixture obtained solvent was evaporated, the evaporation residue obtained was dissolved in DCM and the mixture obtained was extracted with saturated aqueous NaHCO 3 solution.
  • Example 8 Step A 75 mg of the free base from Example 8 Step A was dissolved in 5 mL of DCM and 1 mL of 1 M HCl in diethylether was added. After 10 min the solvent was evaporated and the hydrochlorides of the Example 8 title compounds were obtained in a quantitative manner.
  • Example 9 Step B 100 mg of the free base from Example 9 Step B was dissolved in 5 mL of DCM and 1 mL of 1 M HCl in dietheylether was added. After 10 min the solvent was evaporated and the hydrochlorides of the Example 10 title compounds were obtained in quantitative yield.
  • Example 11 Step A products was obtained in the form of a white foam.
  • Example 12 Step A product in 20 mL of DCM was added 980 mg of mCPBA (70%) and 350 mg of NaHCO 3 .
  • the resulting suspension was stirred at rt overnight, then charged with 20% sodium thiosulfate and extracted with EtOAc.
  • the combined organic layers obtained were washed with saturated aqueous NaHCO 3 , water and brine, dried over sodium sulfate and filtered.
  • the filtrate obtained was concentrated and dried to obtain crude epoxide in the form of a yellowish solid in quantitiative yield.
  • Example 12 Step D products in the form of a colorless foam in quantitative yield.
  • MS-ESI+ (m/z): 522 (M +).
  • Example 13 Step A To a solution of 21.64 g of Example 13 Step A products in 150 mL of DCM was added 22 mL of Boc-anhydride and 25.3 mL of TEA and the resulting reaction mixture was stirred at rt overnight. From the mixture obtained solvent was evaporated and the evaporation residue obtained was charged with EtOAc. The mixture obtained was washed with 1 M HC1, the organic layer obtained was dried over sodium sulfate and the solvent was evaporated to obtain 30.46 g of Example 13 Step B products in the form of a yellow solid.
  • Example 13 Step E products in the form of a colorless foam.
  • Example 13 Step F products To a solution of 300 mg of free base Example 13 Step F products in 1 mL of dioxane was added 0.9 mL of 1 M HCI. After stirring at rt for 1 minutes the solution obtained was lyophilized to obtain 301 mg of the Example 13 title compounds in the form of a colorless foam.
  • Example 15 Step A products from above in 3 mL of DCM was added 0.3 mL of TFA and the mixture obtained was stirred for 6 h at rt.
  • EtOAc and 1 M NaOH were added, the phases obtained were separated and the organic phase obtained was extracted with EtOAc and the combined organic layers obtained were washed with water and brine, dried and the solvent was evaporated.
  • Example 15 Step B products was obtained in the form of a colorless foam.
  • Example 16 Step A products in the form of a colorless solid.
  • Example 16 Step B 7.05 g of mCPBA (70%) was added at 0 °C to a solution of 7.14 g of Example 16 Step B products in 100 ml of DCM.
  • 3.6 g of NaHCO 3 was added and the mixture obtained was stirred at rt for 3 days.
  • the mixture obtained was charged with 20% sodium thiosulfate, stirred for 20 min and extracted with EtOAc.
  • the combined organic layers obtained were washed with saturated aqueous NaHCO 3 solution, water and brine, dried over sodium sulfate and solvent was evaporated.
  • the crude evaporation residue obtained was dried under HV to obtain 5.73 g of Example 16 Step C products in the form of a colorless solid.
  • Example 16 Step D products in 50 mL of DCM was added at 0 °C 520 mg of DTT and 0.94 mL of hydrazine monohydrate and the solution obtained was stirred for 6 h at rt. After addition of 1 M H 3 PO 4 the mixture obtained was diluted with DCM and the layers obtained were separated. The organic layer obtained was washed 3 times with 1 M H 3 PO 4 , once with 1% NaCl solution, dried over sodium sulfate and solvent was evaporated.
  • Example 16 Step E products from above in 30 mL of DCM was added at 0 °C 6.9 mL of TFA.
  • the resulting mixture was stirred for 6 h at rt, charged with EtOAc and saturated aqueous NaHCO 3 solution.
  • the aqueous phase obtained was extracted with EtOAc and the combined organic layers obtained were washed twice with water and brine, dried and solvent was evaporated.
  • Example 16 Step F products To a solution of 3.01 g of free base Example 16 Step F products in 10 mL of dioxane was added 8.06 mL of 1 M HCl. After stirring at rt for 10 minutes the solution obtained was lyophilized to obtain the Example 16 title compounds in the form of a colorless foam in quantitative yield.
  • Example 17 Step A products in 50 ml of THF was added 2.14 g of potassium tert-butoxide and the resulting mixture was stirred at rt for 3 days.
  • the mixture obtained was charged with EtOAc and washed with brine/water, dried over sodium sulfate and solvent was evaporated to obtain 4.3 g of crude Example 17 Step B products in the form of a yellow oil.
  • Example 17 Step B products To a solution of 2 g of Example 17 Step B products in 50 ml of DCM was added 2.98 g of mCPBA (70%) and the resulting mixture was stirred at rt overnight. The mixture obtained was diluted with DCM, washed with 20% sodium thiosulfate solution and brine, dried over sodium sulfate and solvent was evaporated to obtain the crude epoxides as a residue. That residue was taken up in dry toluene and 2.85 mL of thiobenzoic acid and 107 mg of Bu 4 NCl were added and the resulting mixture was stirred at rt overnight.
  • Example 17 Step C early eluting products in 3 mL of DCM was added at 0 °C 20 mg of DTT and 85 ⁇ L of hydrazine monohydrate and the solution obtained was stirred for 6 h at rt. To the mixture obtained further 20 mg of DTT and 85 ⁇ L of hydrazine monohydrate were added and the mixture obtained was stirred for additional 2 h. After addition of 1 M H 3 PO 4 the mixture obtained was diluted with DCM and the layers were separated. The organic layer obtained was washed with brine, dried over magnesium sulfate and solvent was evaporated.
  • Step C late eluting products from Example 17 Step C was treated according to the method described for the early eluting Example 17 Step C products to obtain 35 mg of the Example 17 title compounds in the form of a white solid.

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Abstract

Compounds selected from the group ofN-unsubstituted or N-alkylated or N-acylated 14-O-[(amino(C0-4)alkyl-hydroxy-cycloalkyl- or bicycloalkylsulfanyl)-acetyl]-mutilins which are 14-O-[(amino(C0-4)alkyl-hydroxy-cyclobutylsulfanyl)-acetyl]-mutilins, 14-O-[(amino(C0-4)alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins, 14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins, 14-O-[(amino(C0-4)alkyl-hydroxy-cyclooctylsulfanyl)-acetyl]-mutilins, or 14-O-[(amino(C0-4)alkyt-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins, optionally in the form of a salt and/or a solvate, a pharmaceutical compositions comprising such compounds and their use as pharmaceuticals, e.g. for the treatment of microbial infections and for the treatment of acne, optionally in combination with other pharmaceutically active agents.

Description

  • The present invention relates to organic compounds, namely pleuromutilins.
  • Pleuromutilin, a compound of formula
    Figure imgb0001
     is a naturally occurring antibiotic, e.g. produced by the basidomycetes Pleurotus mutilus and P. passeckerianus, see e.g. The Merck Index, 13th edition, item 7617.
  • A number of further pleuromutilins having the principle ring structure of pleuromutilin and being substituted at the primary hydroxy group have been developed, e.g. as antimicrobials. From WO 2008/113089 pleuromutilin derivatives, e.g. 14-O-[(amino-hydroxy-cyclohexylsulfanyl)-acetyl]-mutilins and from WO 2010/025482 e.g. N-acylated 14-O-[(amino-hydroxy-cyclohexylsulfanyl)-acetyl]-mutilins, are known.
  • We have now found pleuromutilins with interesting antimicrobial activity.
  • In one aspect the present invention provides compounds selected from the group of N-unsubstituted, or N-alkylated, or N-acylated 14-O-[(amino(C0-4)alkyl-hydroxy-cycloalkyl-and bicycloalkylsulfanyl)-acetyl]-mutilins which are
    • 14-O-[(amino(C0-4)alkyl-hydroxy-cyclobutylsulfanyl)-acetyl]-mutilins,
    • 14-O-[(amino(C0-4)alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins,
    • 14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins,
    • 14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins,or
    • 14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins.
  • The term "amino(C0-4)alkyl-" as used herein means that the amino group is either bound to alkyl of 1 to 4 C-atoms or, in case of C0, directly to the (bi)cycloalkylring
  • In one particular aspect the pesent invention provides 14-O-[(amino(C0-4)alkyl-hydroxy-cyclobutylsulfanyl)-acetyl]-mutilins and/or 14-O-[(amino(C0-4)alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins.
  • In another particular aspect the pesent invention provides 14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins and/or 14-O-[(amino(C0-4)-hydroxy-cyclooctylsulfanyl)-acetyl]-mutilins.
  • In a further particular aspect the pesent invention provides 14-O-[amino(C0-4)alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins and/or 14-O-[amino(C0-4)alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins, e.g. 14-O-[amino(C0-4)alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins wherein one ring of the bicyclyic alkyl is a cyclopentyl ring, such as 14-O-[amino(C0-4)alkyl-hydroxy-bicyclo[3.2.0]heptylsulfanyl)-acetyl]-mutilins and 14-O-[amino(C0-4)alkyl-hydroxy-octahydropentalensulfanyl]-acetyl}-mutilins.
  • In a further particular aspect the pesent invention provides 14-O-[(amino(C0-4)alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins, e.g. 14-O-[(amino(C0-4)alkyl -hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins wherein one ring of the bicyclyic alkyl is a cyclopentyl ring, such as 14-O-[(amino(C0-4)alkyl-hydroxy-bicyclo[3.2.0]heptytsutfanyl)-acetyl]-mutilins and 14-O-[(amino(C0-1)alkyl-hydroxy-octahydropentalensulfanyl]-acetyl}-mutilins.
  • In an amino(C0-4)alkyl-group as indicated herein alkyl may be not present for the case of C0; or may be C1alkyl; or may be C2alkyl, or may be C3alkyl, or may be C4alkyl.
  • In another aspect the present invention provides a compound of formula (I)
    Figure imgb0002
     or of formula (II)
    Figure imgb0003
     wherein
    • m is 0, 1, 2, 3 or 4;
    • n is 0, 1, 3 or 4;
    • o is 0, 1, 2, 3 or 4;
    • p is 0, 1, 2, 3 or 4;
    • R is ethyl or vinyl;
    • R1 is hydrogen or (C1-6)alkyl,
    • R2 is hydrogen or
    • - (C3-6)cycloalkyl, or
    • - unsubstituted (C1-6)alkyl, or
    • - (C1-6)alkyl substituted by one or more of
      • hydroxy; preferably one or two,
      • methoxy,
      • halogen,
      • (C3-6)cycloalkyl, or
    • R1 and R2 together with the nitrogen atom to which they are attached form a 5 to 7 membered heterocyclic ring containing at least 1 nitrogen atom or 1 nitrogen and 1 additional heteroatom e. g. selected from N or O, or R2 is a group of formula
      Figure imgb0004
       wherein
    • R3 is hydrogen, straight chain or branched (C1-8)alkyl or (C3-8)cycloalkyl, or
    • R3 is that part of an natural amino acid in D or in L form which remains if the carboxylic acid group is split off, or
    • R3 is that part of a non natural amino acid in D or in L form which remains if the carboxylic acid group is split off.
  • Preferred compounds of the present invention are compounds of formula (III)
    Figure imgb0005
     or of formula (IV)
    Figure imgb0006
     wherein m, n, o, p, R1 and R2 are as defined above, e.g. o is 1; e.g. p is 1 or 2.
  • More preferred compounds of the present invention are compounds of formula (V)
    Figure imgb0007
     or of formula (VI)
    Figure imgb0008
     or of formula (VII)
    Figure imgb0009
     or of formula (VIII)
    Figure imgb0010
     wherein m, R1 and R2 are as defined as defined above.
  • In another aspect the present invention provides a compound of Examples 1 to 17 below, such as a compound of formula I or II, e.g. a compound selected from the group consting of
    • 14-O-{[(1R, 2R, 4R)-4-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof,
    • 14-O-{[(1R, 2R, 4S)-4-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 4R)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof,
    • 14-O-{[(1R, 2R, 4S)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 4S)-4-[(2,2-Dimethyl-propylamino)-methyl]-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 4S)-2-Hydroxy-4-[(2,2,2-trifluoro-acetylamino)-methyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 3S)-2-Hydroxy 3-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 3S)-3-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer thereof,
    • 14-O-{[(1R, 2R, 5R)-5-Hydroxy-2-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    • 14-O-{[(1R, 2R, SR)-2-(2-Amino-ethyl)-5-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    • 14-O-{[(1R, 2R, 5R)-2-[2-(2,2-Dimethyl-propylamino)-ethyl]-5-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    • 14-O-{[(1R, 2R, 5R)-5-Amino-2-hydroxy-cycloheptylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S), (1R, 2R, 5S)- (1S, 2S, 5R) diastereomers thereof,
    • 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Amino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    • 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Formylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    • 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-(2-Amino-acetylamino)-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    • 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Cyclopropylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyll-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    • 14-O-([4-Acetylamino-1-hydroxy-octahydropentalen-2-ylsulfanyl]-acetyl)-mutilin and the diastereomers thereof,
      and
    • 14-O-{[4-Acetylamino-2-hydroxy-octahydropentalen-1-ylsulfanyl]-acetyl} -mutilin and the diastereomers thereof.
  • A compound provided by the present invention is herein also designated as "compound(s) of (according to) the present invention". A compound of the present invention includes a compound in any form, e.g. in free form, in the form of a salt, in the form of a solvate and in the form of a salt and a solvate.
  • According to another aspect, the present invention provides a compound of the present invention in the form of a salt, e.g. and/or solvate.
  • A salt of a compound of the present invention includes preferably pharmaceutically acceptable salts, although pharmaceutically unacceptable salts are included, e.g. for preparation / isolation / purification purposes.
  • A salt of a compound of the present invention includes a base salt or an acid addition salt. Pharmaceutically acceptable acid addition salts include salts of a compound of the present invention with an acid, e.g. hydrogen fumaric acid, fumaric acid, hydrogen tartaric acid, tartaric acid, hydrogen ethane-1,2-disulfonic acid, ethane-1,2-disulfonic acid, hydrogen maleic acid, maleic acid, hydrogen naphthalin-1,5-disulfonic acid, hydrogen naphthalin-1,5-disulfonic acid, acetic acid, hydrogen malic acid, malic acid, lactic acid, hydrogen succinic acid, succinic acid, salicylic acid, hydrogen azelaic acid, azelaic acid, 2-[(2,6-dichlorophenyl)-amino]-benzene acetic acid, hydrogen sulfuric acid, sulfuric acid, dihydrogen phosphoric acid, hydrogen phosphoric acid, phosphoric acid, dihydrogen citric acid, hydrogen citric acid, citric acid, deuterochloric acid, hydrochloric acid, preferably hydrochloric acid.
  • A compound of the present invention in free form may be converted into a corresponding compound in the form of a salt, and vice versa. A compound of the present invention in free form or in the form of a salt and/or in the form of a solvate may be converted into a corresponding compound in free form or in the form of a salt in non-solvated form, and vice versa.
  • A compound of of the present invention may exist in the form of isomers and mixtures thereof, e.g. optical isomers, diastereoisomers, cis/trans conformers. A compound of the present invention may e.g. contain asymmetric carbon atoms and may thus exist in the form of enantiomers or diastereoisomers and mixtures thereof, e.g. racemates or diastereomeric mixtures. Any asymmetric carbon atom may be present in the (R)-, (S)- or (R,S)-configuration, preferably in the (R)- or (S)-configuration.
  • For example, in a compound of formula I or II the carbon atom of the cycloalkyl ring which is attached to S group, the carbon atom of the cycloalkyl ring which is attached to the OH group, and the carbon atom of the cycloalkyl ring to which the (CH2)mN(R1R2) group is attached, are all asymmetric carbon atoms. Substituents attached to such asymmetric carbon atoms may thus exist in (R) and (S) configuration, including mixtures thereof. For example, if in a compound of formula I or II R2 is substituted alkyl and that substituent is attached to a carbon atom of the side chain of such alkyl, the carbon atom to which such substituent is attached is an asymmetric carbon atom and such substituent may be in the (R)- and (S)-configuration, including mixtures thereof.
  • The configuration of asymmetric carbon atoms of the mutilin-tricyclus is preferably the same as in natural pleuromutilin.
  • Isomeric mixtures may be separated as appropriate, e.g. according, e.g. analogously, to a method as conventional, to obtain pure isomers. The present invention includes a compound of the present invention in any isomeric form and in any isomeric mixture. The present invention also includes tautomers of a compound of the present invention, where tautomers can exist.
  • Any compound provided according to the present invention may be obtained as appropriate, e.g. according to, e.g. analogously to a method as conventional or as described herein.
  • E.g. a compound of the present invention may be obtained by reacting a compound PLEU-Tos as defined below with an appropriate amino(C0-4)alkyl-hydroxy-mercapto-cyclobutane, amino(C0-4)alkyl-hydroxy-mercapto-cyclopentane, amino(C0-4)alkyl-hydroxy-mercaptocycloheptane, amino(C0-4)alkyl-hydroxy-mercapto-cyclooctane or amino(C0-4)alkyl-hydroxy-mercapto-bicycloalkane, wherein the amine group is protected, e.g. by Boc, in organic solvent, e.g. polar organic solvent, such as acetonitrile, in the presence of a base and/or coupling agent, such as e.g. DBN; and deprotecting the amine group, e.g. by treatment with an acid, e.g. HC1. The deprotected amine group can be optionally acylated by reaction with a carboxylic acid, e.g. an amino acid, in the presence of a coupling reagent, or with activated carboxylic acid or carboxylic acid derivative, such as an anhydride of the carboxylic acid or carboxylic acid halogenide. The deprotected amine group from above can be optionally alkylated, e.g. by reductive amination or via an alkylating agent.
  • An amino(C0-4)alkyl-hydroxy-mercapto-cyclobutane, amino(C0-4)alkyl-hydroxy-mercaptocyclopentane, amino(C0-4)alkyl-hydroxy-mercapto-cycloheptane amino(C0-4)alkyl-hydroxy-mercapto-cyclooctane, or amino(C0-4)alkyl-hydroxy- mercapto-bicycloalkane,wherein the amine group is protected may be obtained from an N-protected amino(C0-4)alkyl cyclobutene, N-protected amino(C0-4)alkyl cyclopentene, N-protected amino(C0-4)alkyl cycloheptene, N-protected amino(C0-4)alkyl cyclooctene, N-protected amino(C0-4)alkyl bicycloalkene, respectively via epoxidation, ring opening of the epoxide obtained with a thiol e.g. thiobenzoic acid, tritylthiol, thioacetic acid, prefereably with thiobenzoic acid, followed by deprotection of the thiol function e.g. with hydrazine hydrate.
  • The compounds of the present invention exhibit pharmacological activity and are therefore useful as pharmaceuticals.
  • For example, the compounds of the present invention show antimicrobial, e.g. antibacterial, activity against gram positive bacteria, such as coagulase positive Staphylococci, e.g. Staphylococcus aureus, coagulase negative Staphylococci, e.g. Staphylococcus epidermidis, Staphylococcus haemolyticus, and Streptococci, e.g. Streptococcus pyogenes, Streptococcus pneumoniae, Enterococci, e.g. Enterococcus faecium and Listeria monocytogenes and against gram negative bacteria such as Moraxella, e.g. Moraxella catarrhalis, and Haemophilus, e.g. Haemophilus influenzae, and Legionella, e.g. Legionella pneumophila, Neisseriaceae, e.g. Neisseria gonorrhoeae, as well as against Mycoplasms, Chlamydia and obligatory anaerobes, e.g. Bacteroides fragilis, Clostridium difficile, Fusobacterium spp., and Propionibacterium spp.
  • The in vitro activity against aerobic bacteria was determined by Agar Dilution Test or Microdilution Test according to the Clinical and Laboratory Standards Institute (CLSI, former NCCLS) Document M7-A7 Vol.26, No. 2: "Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically - Approved Standard; Seventh Edition (2006)"; and the test against anaerobic bacteria was performed according to the Clinical and Laboratory Standards Institute (CLSI, former NCCLS), Document, M11-A6, Vol. 24, No. 2: "Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria - Approved Standard; Sixth Edition (2004)" and the in vivo activity was tested by the septicaemia mouse model against Staphylococcus aureus.
  • Compounds of the present invention are therefore suitable for the treatment and prevention of diseases which are mediated by microbes, e.g. by bacteria. Diseases which may also be treated include e.g. diseases mediated by Helicobacter, such as Helicobacter pylori, and diseases mediated by Mycobacterium tuberculosis. Diseases which may also be treated include in general inflammatory diseases, where microbes are mediating said inflammation, e.g. including acne.
  • In another aspect the present invention provides a compound of the present invention for use as a pharmaceutical, preferably as an antimicrobial, such as an antibiotic, e.g. an anti-anaerobic.
  • In another aspect the present invention provides a compound of the present invention for use in acne treatment.
  • In a further aspect the present invention provides a compound of the present invention for use in the preparation of a medicament for the treatment of diseases, mediated by microbes, such as bacterials, for example
    • diseases mediated by bacteria, e.g. selected from Staphylococci, Streptococci, Enterococci;
    • diseases mediated by bacteria, e.g. selected from Moraxella, Haemophilus, Legionella, Neisseriaceae;
    • diseases mediated by Helicobacter;
    • diseases mediated by Mycobacterium tuberculosis;
    • e.g. diseases mediated by Mycoplasms, Chlamydia and obligatory anaerobes;
    and for the treatment of acne.
  • In a further aspect the present invention provides a method of treatment of diseases mediated by microbes which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention e.g. in the form of a pharmaceutical composition.
  • In a further aspect the present invention provides a method of treatment of acne which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention e.g. in the form of a pharmaceutical composition.
  • Treatment includes treatment and prophylaxis.
  • For antimicrobial and acne treatment, the appropriate dosage will, of course, vary depending upon, for example, the chemical nature and the pharmacokinetic data of a compound of the present invention employed, the individual host, the mode of administration and the nature and severity of the conditions being treated. However, in general, for satisfactory results in larger mammals, for example humans, an indicated daily dosage is in the range of about 0.5 mg to 3 g of a compound of the present invention conveniently administered, for example, in divided doses up to four times a day.
  • A compound of the present invention may be administered by any conventional route, for example enterally, e.g. including nasal, buccal, rectal, oral administration; parenterally, e.g. including intravenous, intramuscular, subcutaneous administration; or topically, e.g. including epicutaneous, intranasal, intratracheal administration, e.g. in form of coated or uncoated tablets, capsules, injectable solutions or suspensions, e.g. in the form of ampoules, vials, in the form of creams, ointments, gels, pastes, inhaler powder, foams, tinctures, lipsticks, drops, sprays, or in the form of suppositories, e.g. in analogous manner to macrolides, such as erythromycins, e.g. clarithromycin or azithromycin.
  • A compound of the present invention may be administered in the form of a pharmaceutically acceptable salt, e.g. an acid addition salt or a base addition salt, e.g. a metal salt, or in free form, optionally in the form of a solvate. A compound of the present invention in the form of a salt exhibits the same order of activity as the compound in free form, optionally in the form of a solvate.
  • A compound of the present invention may be used for pharmaceutical treatment according to the present invention alone or in combination with one or more other pharmaceutically active agents. Such other pharmaceutically active agents include e.g. other antibiotics and antiinflammatory agents, and, if a compound of the present invention is used in the treatment of acne, other pharmaceutically agents include furthermore agents which are active against acne.
  • Combinations include fixed combinations, in which two or more pharmaceutically active agents are in the same formulation; kits, in which two or more pharmaceutically active agents in separate formulations are sold in the same package, e.g. with instruction for coadministration; and free combinations in which the pharmaceutically active agents are packaged separately, but instruction for simultaneous or sequential administration are given.
  • In another aspect the present invention provides a pharmaceutical composition comprising a compound of the present invention in free form or in the form of a pharmaceutically acceptable salt and/or in the form of a solvate as such or in association with at least one pharmaceutical, excipient, e.g. carrier or diluent, e.g. including fillers, binders, disintegrators, flow conditioners, lubricants, sugars and sweeteners, fragrances, preservatives, stabilizers, wetting agents and/or emulsifiers, solubilizers, salts for regulating osmotic pressure and/or buffers.
  • In another aspect the present invention provides a pharmaceutical composition according to the present invention, further comprising another pharmaceutically active agent.
  • Such pharmaceutical compositions may be manufactured according, e.g. analogously, to a method as conventional, e.g. by mixing, granulating, coating, dissolving or lyophilizing processes. Unit dosage form may contain, for example, from about 0.5 mg to about 2000 mg, such as 10 mg to about 500 mg.
  • The compounds of the present invention are additionally suitable as veterinary agents, e.g. veterinary active compounds, e.g. in the prophylaxis and in the treatment of microbial, e.g. bacterial diseases, in animals, such as fowl, pigs and calves, e.g., and for diluting fluids for artificial insemination and for egg-dipping techniques.
  • In another aspect the present invention provides a compound of the present invention for use as a veterinary agent.
  • In a further aspect the present invention provides a compound of the present invention for the preparation of a veterinary composition which is useful as a veterinary agent.
  • In another aspect the present invention provides a veterinary method for the prophylaxis and the treatment of microbial, e.g. bacterial diseases which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention, e.g. in the form of a veterinary composition.
  • The compounds of Examples 1 to 17 following thereafter exhibit MICs ≤ 2 µg/mL against Staphylococcus aureus ATCC49951 and Streptococcus pneumoniae ATCC49619.
  • The trivial name mutilin refers to the IUPAC systematic name (1S, 2R, 3S, 4S, 6R, 7R, 8R, 14R)-3,6-dihydroxy-2,4,7,14-tetramethyl-4-vinyl-tricyclo[5.4.3.01, 8]tetradecan-9-one. In the examples, pleuromutilin derivatives are numbered in analogy to the mutilin numbering system described by H. Berner (Berner, H.; Schulz, G.; Schneider H. Tetrahedron 1980, 36, 1807-1811.):
    Figure imgb0011
  • Herein, including the examples the following abbreviations are used:
    • °C
    degree Celsius
    1H NMR
    proton nuclear magnetic resonance spectroscopy
    Boc,boc
    tert-butoxycarbonyl
    Bu4NC1
    tetrabutylammonium chloride
    CH
    cyclohexane
    d
    day(s)
    DBN
    1,5-diazobicyclo[4.3.0]non-5-ene
    DCM
    dichloromethane
    DIPE
    diisopropylether
    DMAP
    4-(dimethylamino)-pyridine
    DMF
    N,N-dimethylformamide
    DMSO
    dimethylsulfoxide
    DTT
    1,4-dithio-DL-threitol
    EDC
    N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride
    ESI+
    electrospray ionization in positive mode
    ESI-
    electrospray ionization in negative mode
    EtOAc
    ethyl acetate
    h
    hour(s)
    HCl
    hydrochloric acid
    HOBT
    1-hydroxybenzotriazole
    HV
    high vacuum
    LAH
    lithium aluminium hydride
    M
    molarity
    mCPBA
    metachloroperoxybenzoic acid
    MeOH
    methanol
    MIC
    minimum inhibitory concentration
    min
    minutes
    MS
    mass spectrometry
    MTBE
    methyl tert-butyl ether
    m/z
    mass/charge ratio
    Ph
    phenyl
    Rf
    retention factor, retardation factor
    rt
    room temperature
    TLC
    thin layer chromatography
    TEA, Et3N
    triethylamine
    TFA
    trifluoroacetic acid
    THF
    tetrahydrofuran
  • PLEU-Tos is a compound of formula
    Figure imgb0012
  • PLEU is a residue of formula
    Figure imgb0013
  • In the following examples all temperatures are in degrees Celsius (°C) and are uncorrected.
    • Example 1 14-O-{[(1R, 2R, 4R)-4-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4S) diastereomer thereof A. syn and anti tert-Butyl (6-Oxa-bicyclo[3.1.0]hex-3-yl)-carbamate
  • Figure imgb0014
  • 6.63 g of mCPBA was added at 0°C to a solution of 4.7 g of N-Boc-cyclopent-3-enyl amine in 80 mL of DCM. The solution obtained was stirred at rt for 1.5 h, charged with 10 mL of 20% sodium thiosulfate, stirred for 10 min, then DCM was added and the organic layer obtained was washed with saturated NaHCO3 solution. The aqueous layer obtained was extracted with DCM and the combined organic layers were dried over magnesium sulfate and the solvent was evaporated. 5.2 g of product was obtained which was recrystallised twice from heptane. After crystallization, 1.01 g of syn epoxide was obtained in the form of white needles. The mother liquors were subjected to chromatography (silica, toluene/EtOAc = 9:1 - 7:1) to yield 3.51 g of anti epoxide in the form of a white amorphous solid.
  • Syn epoxide: is NMR (400MHz, DMSO-d6, δ, ppm, characteristic signals): 1.36 (s, 9H), 6.79 (d, 1H, J=8Hz), 1.47 (dd, 1H, J=8Hz and 14Hz), 2.17 (dd, 1H, J=8Hz and 14Hz), 3.44 (s, 2H), 3.54 (m, 1H). MS-ESI+ (m/z): 200 (M + H). TLC: toluene/EtOAc = 3:1, Rf = 0.4.
  • Anti epoxide: 1H NMR (400MHz, DMSO-d6, δ, ppm, characteristic signals): 1.35 (s, 9H), 1.78 (d, 2H, J=15Hz), 1.98 (dd, 1H, J=8Hz and 15Hz), 3.51 (s, 2H), 3.89 (q, 1H, J=8 and 16Hz), 5.70 (d, 1H, J=8Hz). MS-ESI+ (m/z): 200 (M + H). TLC: toluene/EtOAc = 3:1, Rf = 0.45.
    • B. Thiobenzoic acid S-((1R, 2R, 4R)-4-tert-butoxycarbonylamino-2-hydroxy-cyclopentyl) ester and the (1S, 2S, 4S) diastereomer thereof
  • Figure imgb0015
  • 9.88 mL of thiobenzoic acid was added dropwise to a solution of 2.81 g of the anti epoxide from Example 1 Step A in 50 mL of toluene. To the mixture obtained 248 mg of Bu4NCl was-added and the resulting solution was stirred for 16 h at rt. After this time further 2 mL of thiobenzoic acid together with 500 mg of Bu4NCl was added and stirring was continued for another 22 h. The mixture obtained was charged with saturated NaHCO3 solution and stirred for 15 min. To the mixture obtained EtOAc was added and the organic phase was washed 4 times with saturated NaHCO3 solution, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 98:2 - 8:1) to yield 4.65 g of Example 1 Step B products in the form of a white solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 1.38 (s, 9H), 1.48 (m, 1H), 1.77 (m, 1H), 2.20 (m, 2H), 3.68 - 4.02 (m, 3H), 5.27 (d, 1H, J=6Hz), 6.98 (bd, 1H, J=8Hz), 7.40 - 7.92 (m, 5H). MS-ESI+ (m/z): 338 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.3.
    • C. 14-0-{[(1R, 2R, 4R)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer
  • Figure imgb0016
  • To a solution of 600 mg of Example 1 Step B products in 3 mL of DCM was added 180 µL of hydrazine monohydrate in two portions and the solution obtained was stirred for 20 min at rt. After addition of 4 mL of 1 M HC1 the mixture obtained was stirred for 5 min, diluted with DCM and the layers were separated. The organic layer obtained was dried and the solvent was evaporated. The evaporation residue obtained was dissolved in 10 mL of acetonitrile. 219 µL of DBN and 799 mg of pleuromutilin tosylate were added and the solution obtained was stirred for 1 h at rt. EtOAc was added and the organic phase was washed with brine, dried and the solvent was evaporated. The residue was subjected to chromatography (silica, toluene/EtOAc = 3:1 - 1:1) to obtain 797 mg of Example 1 Step C products in the form of white foam.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 0.62 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.14 (s, 3H), 1.37 (s, 12H), 2.42 (bs, 1H), 3.06 (m, 1H), 3.36 (m, 3H), 3.77 (m, 2H), 4.53 (d, 1H, J=6Hz), 5.08 (m, 3H), 5.54 (bd, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 6.82 (t, 1H, J=4Hz). MS-ESI+ (m/z): 594 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.45.
    • D. 14-O-{[(1R, 2R, 4R)-4-tert-Amino-2-hvdroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4S) diastereomer
  • Figure imgb0017
  • 780 mg of Example 1 Step C products was dissolved in 10 mL of DCM, 8 mL of 1 M HC1 in diethylether was added and the solution stirred for 16 h at rt. Further 2 mL of 1 M HC1 in diethylether was added and stirred for another 8 h. After evaporation of the solvent the solid residue was triturated with diethylether. After filtration, 579 mg of the Example 1 Step D products was obtained as white solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.42 (bs, 1H), 3.12 - 3.60 (m, 5H), 3.90 (m, 1H), 4.59 (bs, 1H), 5.06 (m, 2H), 5.53 (bs, 1H), 5.60 (bd, 1H, J=12Hz), 6.13 (dd, 1H, J=11 and 18Hz), 8.06 (bs, 3H). MS-ESI+ (m/z): 494 (M + H). TLC: DCM/MeOH = 6:1+1% NH4OH, Rf = 0.5.
    • Example 2 14-O-{[(1R, 2R, 4S)-4-Amino-2-hydroxy-cyclopentylsulfanyl)-acetyl}-mutilin hydrochloride and the (1S, 2S, 4R) diastereomer thereof A. Thiobenzoic acid S-((1R, 2R, 4S)-4-tert-butoxycarbonylamino-2-hydroxy-cyclopentyl) ester and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0018
  • 1.04 g of the syn epoxide from Example 1 Step A was treated according to the method of Example 1 Step B to obtain after chromatography (silica, toluene/EtOAc = 98:2 - 7:1) 1.51 g of Example 2 Step A products as a white solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 1.37 (s, 9H), 1.45 (m, 1H), 1.80 (m, 2H), 2.55 (m, 1H), 3.60 (m, 1H), 4.02 (m, 2H), 5.19 (d, 1H, J=4Hz), 6.98 (d, 1H, J=6Hz), 7.40 - 7.90 (m, 5H). MS-ESI+ (m/z): 675 (2M + H), 338 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.25.
    • B. 14-O-{[(1R, 2R, 4S)-4-tert-Butoxycarbonylamino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer
  • Figure imgb0019
  • 400 mg of Example 2 Step A products was treated according to the method of Example 1 Step C to obtain after chromatography (silica, toluene/EtOAc = 2:1 - 1:2) 510 mg of Example 2 Step B products as a white solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 0.62 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.12 (s, 3H), 1.36 (s, 12H), 2.42 (bs, 1H), 2.89 (m, 1H), 3.40 (m, 3H), 3.90 (m, 1H), 4.54 (d, 1H, J=6Hz), 4.99 (d, 1H, J=4Hz), 5.08 (m, 2H), 5.54 (bd, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 6.88 (bd, 1H, J=8Hz). MS-ESI+ (m/z): 594 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.4.
    • C. 14-O-{[(1R, 2R, 4S)-4-tert-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4R) diastereomer
  • Figure imgb0020
  • 490 mg of the Example 2 Step B products was treated according to the method of Example 1 Step C to obtain 405 mg of Example 2 Step C products as a white solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.42 (bs, 1H), 2.97 (m, 1H), 3.43 (m, 3H), 4.00 (m, 1H), 4.58 (d, 1H, J=6Hz), 5.08 (m, 2H), 5.26 (d, 1H, J=4Hz), 5.55 (bd, 1H, J=9Hz), 6.13 (dd, 1H, J=11 and 18Hz), 8.10 (bs, 3H). MS-ESI+ (m/z): 494 (M + H). TLC: DCM/MeOH = 6:1 + 1% NH4OH, Rf = 0.5.
    • Example 3 14-O-{[(1R, 2R, 4R)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4S) diastereomer thereof A. Cyclopent-3-enyl-methanol
  • Figure imgb0021
  • 3.66g of LAH was suspended in 100 mL of THF and 10.49 g of cyclopent-3-enecarboxylic acid dissolved in 40 mL of THF was added dropwise. The mixture was kept at reflux for 2 h, cooled to rt and slowly water and 10% sulfuric acid were added. The mixture obtained was extracted with diethylether and the organic phase dried and carefully evaporated using a cold water bath. 10 g of Example 3 Step A product (still contains THF) was obtained as colorless liquid.
  • 1H-NMR (200 MHz, CDC13, δ, ppm, characteristic signals): 1.73 (s, 1H), 2.10 (m, 2H), 2.50 (m, 3H), 3.49 (d, 2H, J=16Hz), 5.66 (s, 2H). TLC: toluene/EtOAc = 3:1, Rf = 0.35.
    • B. Methanesulfonic acid cyclopent-3-enylmethyl ester
  • Figure imgb0022
  • 10.75 g of cyclopent-3-enyl-methanol from Example 3 Step A was dissolved in 200 mL of THF and 21 mL of N-methylmorpholine was added. 25.26 g of methanesulfonic anhydride was added in portions and the reaction mixture was stirred for 20 h at rt. EtOAc was added and the organic phase washed with saturated NaHCO3 solution and 1 M HC1. The organic phase was dried and evaporated. 18.37g of Example 3 Step B product was obtained as slightly yellow oil.
  • 1H-NMR (200 MHz, CDC13, δ, ppm, characteristic signals): 2.12 (m, 2H), 2.55 (m, 2H), 2.70 (m, 1H), 3.01 (s, 3H), 4.12 (d, 2H, J=8Hz), 5.66 (s, 2H). MS-ESI+ (m/z): 375 (2M + Na), 199 (M + Na). TLC: toluene /EtOAc = 3:1, Rf = 0.5.
    • C. tert-Butyl Cyclopent-3-enylmethyl-carbamate
  • Figure imgb0023
  • To 18.37g of Example 3 Step B product dissolved in 200 mL of DMF was added 27.1 g of sodium azide and the reaction mixture obtained was stirred at 60 °C for 20 h. To the mixture obtained diethylether was added, the organic layer obtained was washed with water, dried and carefully evaporated using a cold water bath. The crude azide product still contains DMF and is used for the next step without further purification. 18.6 g of this crude azide product was dissolved in 150 mL of THF and 5 mL of water. To the mixture obtained 26.17 g of triphenylphosphine was added and the reaction mixture obtained was stirred at 80 °C for 4 h. The mixture obtained was allowed to cool down to 4 °C using an ice bath and 18.3 g of Boc-anhydride dissolved in 50 mL of THF was added. The mixture obtained was stirred at rt for 1.5 h, the solvent was evaporated and the evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 99:1 - 95:5) to obtain 13.16 g of Example 3 Step C product in the form of a colorless oil.
  • 1H-NMR (200 MHz, CDC13, δ, ppm, characteristic signals): 1.44 (s, 9H), 2.03 (m, 2H), 2.45 (m, 3H), 3.09 (bs, 2H), 5.65 (s, 2H). MS-ESI+ (m/z): 395 (2M + H), 142 (M - tBu). TLC: toluene/EtOAc = 9:1, Rf = 0.5.
    • D. Syn and anti tert-Butyl (6-Oxa-bicyclo[3.1.0]hex-3-ylmethyl)-carbamate
  • Figure imgb0024
  • 19.17 g of mCPBA was added at 0°C to a solution of 14.61 g of Example 3 Step C product in 130 mL of DCM. The solution obtained was stirred at rt for 1 h, poured onto 20% sodium thiosulfate and the mixture obtained was stirred for 10 min. To the mixture obtained DCM was added, the phases obtained were separated and the organic layer obtained was washed with saturated NaHCO3 solution. The aqueous layer obtained was extracted with DCM and the combined organic layers were dried over MgSO4 and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 95:5 - 8: 1) and the fractions obtained were recrystallised from EtOAc/mixed heptanes and again subjected to chromatography (silica, CH/EtOAc = 9:1 - 6: 1) to obtain overall 4.21 g of anti epoxide and 10.03 g of syn epoxide of Example 3 Step D product in the form of a white solids.
  • Anti epoxide: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.36 (s, 9H), 1.70 (m, 4H), 2.05 (m, 1H), 2.75 (dd, 2H, J=6Hz and 8Hz), 3.44 (s, 2H), 6.83 (t, 2H, J=6Hz). MS-ESI+ (m/z): 427 (2M + H), 214 (M + H), 158 (M - tBu). TLC: CH/EtOAc = 8:1, Rf = 0.55.
  • Syn epoxide: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.28 (m, 2H), 1.37 (s, 9H), 1.73 (m, 1H), 1.94 (m, 2H), 2.89 (t, 2H, J=6Hz), 3.41 (s, 2H), 6.84 (t, 1H, J=8Hz). MS-ESI+ (m/z): 449 (2M + Na), 214 (M + H), 158 (M - tBu). TLC: CH/EtOAc = 8:1, Rf = 0.45.
    • E. Thiobenzoic acid S-[(1R, 2R, 4R)-4-(tert-butoxycarbonylamino-methyl)-2-hydroxycyclopentyl] ester and the (1S, 2S, 4S) diastereomer thereof
  • Figure imgb0025
  • 18.2 mL of thiobenzoic acid was added dropwise to a solution of 4.18 g of anti epoxide from Example 3 Step D in 110 mL of toluene containing 461 mg of Bu4NCl. The reaction mixture obtained was stirred for 20 h at rt. To the mixture obtained saturated NaHCO3 solution, and after 10 min, EtOAc was added, the phases obtained were separated and the organic phase obtained was washed 4 times with saturated NaHCO3 solution, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 97:3 - 9:1) to yield 5.79 g of Example 3 Step E products in the form of a colorless oil.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.26 (m, 1H), 1.36 (s, 9H), 1.70 (m, 1H), 2.05 (m, 2H), 2.18 (m, 1H), 2.96 (m, 2H), 3.70 (m, 1H), 4.00 (m, 1H), 5.18 (d, 1H, J=4Hz), 6.90 (t, 1H, J=6Hz), 7.45 - 7.92 (m, 5H). MS-ESI+ (m/z): 352 (M + H), 296 (M - tBu). TLC: DCM/MeOH = 95:5, Rf = 0.35.
    • F. 14-O-[(1R, 2R, 4R)-4-tert-Butoxycarbonylaminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof
  • Figure imgb0026
  • To a solution of 5.53 g of Example 3 Step E products in 80 mL of acetonitrile was added 1.56 mL of hydrazine monohydrate and the solution obtained was stirred for 30 min at rt. After addition of 25 mL of 1 M HCl the mixture obtained was stirred for 5 min, diluted with DCM and the layers were separated. The organic layer obtained was dried and solvent was evaporated. The evaporation residue obtained was dissolved in 100 mL of acetonitrile. 2.06 mL of DBN and 7.53 g of pleuromutilin tosylate was added and the solution obtained was stirred for 4 h at rt. To the mixture obtained EtOAc was added, the phases obtained were separated and the organic phase obtained was washed with brine, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica using DCM/MeOH = 100:2) to yield 7.16 g of Example 3 Step F products in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.41 (bs, 1H), 2.87 (t, 2H, J=6Hz), 2.99 (m, 1H), 3.45 (m, 3H), 3.83 (m, 1H), 4.53 (d, 1H, J=6Hz), 4.98 (d, 1H, J=4Hz), 5.08 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 6.84 (t, 1H, J=6Hz). MS-ESI+ (m/z): 608 (M + H). TLC: DCM/MeOH = 95:5, Rf= 0.35.
    • G. 14-O-{[(1R, 2R, 4R)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof
  • Figure imgb0027
  • 1g of Example 3 Step F products was dissolved in 20 mL of DCM and 5 mL of 2 M HC1 in diethylether was added. After 16 h at rt the solvent was evaporated and the evaporation residue obtained was subjected to chromatography (silica, DCM/MeOH/NH4OH 100:8:0.8) to obtain 155 mg of the free base Example 3 Step G products in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.42 (bs, 1H), 3.04 (m, 2H), 3.30 - 3.45 (m, 4H), 3.83 (m, 1H), 4.54 (d, 1H, J=6Hz), 5.06 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz). MS-ESI+ (m/z): 508 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.15.
    • H. 14-O-{[(1R, 2R, 4R)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4S) diastereomer thereof
  • Figure imgb0028
  • 150 mg of the free base from Example 3 Step G was dissolved in 5 mL of DCM. and 1 mL of 1 M HCl in diethylether was added. After 10 min the solvent was evaporated and the hydrochloride Example 3 Step H products were obtained in the form of a white solid in a quantitative manner.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.42 (bs, 1H), 2.75 (m, 1H), 3.10 (m, 1H), 3.30 - 3.45 (m, 3H), 3.59 (d, 1H, J=8Hz), 3.92 (m, 1H), 4.57 (d, 1H, J=6Hz), 5.08 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 7.87 (bs, 3H). MS-ESI+ (m/z): 508 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.15.
    • Example 4 14-O-{[(1R, 2R, 4S)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4R) diastereomer thereof A. Thiobenzoic acid S-[(1R, 2R, 4S)-4-(tert-butoxycarbonylamino-methyl)-2-hydroxycyclopentyl] ester and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0029
  • 5 g of the syn epoxide from Example 3 Step D was treated according to the method of Example 3 Step E to obtain after chromatography (silica, toluene/EtOAc = 9:1-7:1) 7.11 g of Example 4 Step A products in the form of a amorphous white solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.27 (m, 1H), 1.37 (s, 9H), 1.62 (m, 2H), 2.30 (m, 2H), 2.89 (m, 2H), 3.63 (m, 1H), 4.00 (m, 1H), 5.11 (d, 1H, J=4Hz), 6.92 (t, 1H, J=6Hz), 7.50-7.95 (m, 5H). MS-ESI+ (m/z): 752 (2M + Na), 703 (2M + H), 352 (M + H), 296 (M - tBu). TLC: DCM/MeOH = 95:5, Rf = 0.4.
    • B. 14-O-{[(1R, 2R, 4S)-4-tert-Butoxycarbonylaminomethyl-2-hydroxy-cvclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0030
  • 7 g of the Example 4 Step A products was treated according to the method of Example 3 Step F to obtain after chromatography (silica, DCM/MeOH = 100:2) 9.41 g of the Example 4 Step B products in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.42 (bs, 1H), 2.84 (t, 2H, J = 6Hz), 2.93 (m, 1H), 3.25 - 3.48 (m, 3H), 3.87 (m, 1H), 4.53 (d, 1H, J=6Hz), 4.89 (d, 1H, J=6Hz), 5.08 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 6.86 (t, 1H, J=4Hz). MS-ESI+ (m/z): 608 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.35.
    • C. 14-O-{(1R, 2R, 4S)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0031
  • 2 g of Example 4 Step B products was treated according to the method of Example 3 Step G to obtain after chromatography (silica, DCM / MeOH / NH4OH 100:8:0.8) 906 mg of the free base Example 4 Step C products.
  • TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.15.
    • D. 14-O-1[(1R, 2R, 4S)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0032
  • 200 mg of Example 4 Step C products was dissolved in 5 mL of DCM and 1 mL of 1 M HC1 in diethylether was added. After 10 min the solvent was evaporated and 166 mg of the hydrochloride Example 4 Step D products was obtained in the form of a white amorphous solid.
  • 1H-NMR (200 MHz, DMSO-db, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.42 (bs, 1H), 2.75 (t, 2H, J=6Hz), 3.01 (m, 1H), 3.40 (m, 3H), 3.95 (m, 1H), 4.56 (d, 1H, J=6Hz), 5.06 (m, 3H), 5.55 (d, 1H, J=6Hz), 6.14 (dd, 1H, J=11 and 18Hz), 7.87 (bs, 3H). MS-ESI+ (m/z): 508 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.15.
    • Example 5 14-O-{[(1R, 2R, 4S)-4-[(2,2-Dimethyl-propylamino)-methyl]-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0033
  • 300 mg of Example 4 Step C products was dissolved in 15 mL of dry DCM containing molecular sieves. To the mixture obtained 42 mg of pivalaldehyde was added at rt and after 1 h the reaction mixture was charged with 136 mg of sodium triacetoxyborohydride and stirring was continued for another 23 h. To the mixture obtained DCM was added, the phases obtained were separated and the organic layer obtained was washed with saturated NaHCO3 solution, dried and evaporated. After chromatography (silica, DCM / MeOH = 95:5 - 9:1) 68 mg of the free base products of the Example 5, which was obtained in the form of a white foam, was dissolved in 3 mL of DCM and 300 µL of 2M HCl in diethylether was added, the mixture obtained was stirred for 10 min and the solvent was evaporated. 52 mg of the Example 5 title compounds in the form of a hydrochloride was obtained in the form of a white solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J = 6Hz), 0.82 (d, 3H, J = 6Hz), 1.06 (s, 3H), 1.07 (s, 9H), 1.45 (s, 3H), 2.42 (bs, 1H), 2.73 (m, 2H), 2.90 (m, 2H), 3.02 (m, 1H), 3.30 ― 3.45 (m, 3H), 3.92 (m, 1H), 4.58 (d, 1H, J=6Hz), 5.08 (m, 3H), 5.55 (d, 1H, J=8Hz), 6.15 (dd, 1H, J=11 and 18Hz), 8.39 (bs, 3H). MS-ESI- (m/z): 622 (M + HCOO-), MS-ESI- (m/z): 578 (M+). TLC: DCM/MeOH = 9:1, Rf = 0.25.
    • Example 6 14-O-{[(1R, 2R, 4S)-2-Hydroxy-4-[(2,2,2-trifluoro-acetylamino)-methyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof
  • Figure imgb0034
  • 2g (3.29 mmol) of Example 4 Step B products was dissolved in 20 mL of DCM and 2 mL of TFA was added and the mixture obtained was stirred for 7 h at rt. To the mixture obtained saturated NaHCO3 solution was added, the phases obtained were separated and the organic phase obtained was extracted with DCM, dried and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, DCM using CH2C12 / MeOH = 98:2) to obtain 70 mg of the Example 6 title compounds in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.41 (bs, 1H), 2.97 (m, 1H), 3.13 (t, 2H, J=6Hz), 3.35 (m, 2H), 3.42 (m, 1H), 3.89 (m, 1H), 4.54 (d, 1H; J=6Hz), 4.95 (d, 1H, J=6Hz), 5.08 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 9.45 (t, 1H, J=6Hz). MS-ESI-(m/z): 648 (M + HCOO-). TLC: DCM/MeOH = 9:1, Rf = 0.65.
    • Example 7 14-O-{[(1R, 2R, 3S)-2-Hydroxy-3-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer thereof A. 2-Cyclopent-2-enyl-ethanol
  • Figure imgb0035
  • 3.1 g of LAH was suspended in 100 mL of THF and 9.55 g of cyclopent-2-enyl-acetic acid dissolved in 40 mL of THF was added dropwise. The mixture obtained was kept at reflux for 2 h, cooled to rt and water and 10% sulfuric acid was added slowly. The mixture obtained was extracted with diethylether, the organic phase obtained was dried and carefully evaporated using a cold water bath to obtain 11 g of Example 7 Step A product (still containing THF) in the form of a colorless liquid.
  • 1H-NMR (200 MHz, CDC13, δ, ppm, characteristic signals): 1.42 (m, 1H), 1.49 (s, 1H), 1.62 (m, 2H), 2.06 (m, 1H), 2.32 (m, 1H), 2.76 (m, 1H), 3.69 (t, 2H, J=6Hz), 5.70 (m, 2H). TLC: toluene/EtOAc = 3:1, Rf = 0.4.
    • B. Methanesulfonic acid 2-cyclopent-2-enyl-ethyl ester
  • Figure imgb0036
  • 8.8 g of cyclopent-3-enyl-ethanol from Example 7 Step A was dissolved in 200 mL of THF and 14.67 mL of N-methylmorpholine was added. To the mixture obtained 17.77 g of methanesulfonic anhydride was added in portions and the reaction mixture obtained was stirred for 20 h at rt. Further 1.5 mL of N-methylmorpholine and 1.8 g of methanesulfonic anhydride, and after another 4 h, EtOAc was added, the phases obtained were separated and the organic phase obtained was washed with saturated NaHCO3 solution and 1 M HC1. The organic phase obtained was dried and the solvent was evaporated to obtain 13.89 g of Example 7 Step B product in the form of a slightly yellow oil.
  • 1H-NMR (200 MHz, CDC13, δ, ppm, characteristic signals): 1.41 (m, 1H), 1.80 (m, 2H), 2.10 (m, 1H), 2.32 (m, 1H), 2.80 (m, 1H), 3.01 (s, 3H), 4.27 (t, 2H, J=6Hz), 5.63 and 5.77 (2xm, 2H). MS-ESI+ (m/z): 403 (2M + Na), 213 (M +Na), 208 (M + NH4). TLC: toluene/EtOAc = 3:1, Rf = 0.65.
    • C. tert-Butyl (2-Cyclopent-2-enyl-ethyl)-carbamate
  • Figure imgb0037
  • To 13.89 g of methanesulfonic acid cyclopent-3-enyl-ethyl ester from Example 7 Step B dissolved in 150 mL of DMF was added 18.9 g of sodium azide and the reaction mixture obtained was stirred at 60°C for 20 h. To the mixture obtained diethylether was added, the phases obtained were separated and the organic layer washed with water, dried and carefully evaporated using a cold water bath. The crude azide product obtained still contains DMF and is used for the next Step without further purification.
  • The 14.14 g of the crude azide form above was dissolved in 130 mL of THF and 5 mL of water. 22.98 g of triphenylphosphine was added and the reaction mixture obtained was stirred at 80 °C for 4 h. The mixture obtained was allowed to cool down to 4 °C using an ice bath and to the mixture obtained 17.5 g of Boc-anhydride dissolved in 50 mL of THF was added. The reaction mixture obtained was stirred at rt for 1.5 h, the solvent was evaporated and the evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 99:1 - 95:5) to obtain 2.05 g of the Example 7 Step C product in the form of a as colorless oil.
  • 'H-NMR (200 MHz, CDCl3, δ, ppm, characteristic signals): 1.25 - 1.62 (m, 3H), 1.37 (s, 9H), 2.02 (m, 1H), 2.25 (m, 1H), 2.62 (m, 1H), 3.11 (t, 2H, J=8Hz), 4.50 (bs, 1H), 5.54 - 5.70 (m, 2H). MS-ESI+ (m/z): 423 (2M + H). TLC: toluene/EtOAc = 9:1, Rf = 0.65.
    • D. Syn-[2-(6-Oxa-bicyclo[3.1.0]hex-2-yl)-ethyl]-carbamic acid tert-butyl ester
  • Figure imgb0038
  • 2.39 g of mCPBA was added at 0 °C to a solution of 1.95 g of Example 7 Step C product in 30 mL of DCM. The solution obtained was stirred at rt for 1.5 h, poured into 20% aqueous sodium thiosulfate solution and the mixture obtained was stirred for 10 min. To the mixture obtained DCM was added, the phases obtained were separated and the organic layer obtained was washed with aqueous saturated NaHCO3 solution. The aqueous layer obtained was extracted with DCM and the combined organic layers obtained were dried over MgSO4 and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 95:5 - 9:1) to yield 1.8 g of Example 7 Step D products in the form of a colorless oil.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.25 - 1.70 (m, 5H), 1.39 (s, 9H), 1.88 (m, 2H), 2.98 (m, 2H), 3.35 and 3.42 (2xm, 2H), 6.87 (bs, 1H). MS-ESI+ (m/z): 477 (2M + Na), 228 (M + H), 172 (M - tBu), 154 (M - OtBu). TLC: toluene/EtoAc = 3:1, Rf = 0.65.
    • E. Thiobenzoic acid S-[(1R, 2R, 5R)-5-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxycyclopentyl) ester and the (1S, 2S, 5S) diastereomer thereof and Thiobenzoic acid S-[(1R, 2R, 3S)-3-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxycyclopentyl) ester and the (1S, 2S, 3R) diastereomer thereof
  • Figure imgb0039
     9.08 mL of thiobenzoic acid was added dropwise to a solution of 1.8 g of syn epoxides from Example 7 Step D in 40 mL of toluene containing 187 mg of Bu4NCl. The reaction mixture was stirred for 20 h at rt. To the mixture obtained saturated NaHCO3 solution was added and after 10 min the reaction mixture was charged with EtOAc and the organic phase obtained was washed 4 times with saturated NaHCO3 solution, dried and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 95:5 ― 8:1) to obtain 1.28 g of thiobenzoic acid S-[(1R,2R,SR)-5-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxy-cyclopentyl] ester and the (1S, 2S, 5S) diasteromer thereof (late eluting compounds) in the form of a slightly yellow oil as well as 875 mg of thiobenzoic acid S-[(1R,2R,3S)-3-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxycyclopentyl] ester and the (1S, 2S, 3R) diasteromer thereof (early eluting compounds) in the form of a colorless oil.
  • Late eluting products: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.30 ― 1.95 (m, 7H), 1.38 (s, 9H), 2.93 (m, 2H), 3.45 (m, 1H), 3.99 (m, 1H), 5.11 (d, 1H, J=6Hz), 6.97 (t, 1H, J=6Hz), 7.50 ― 7.98 (m, 5H). MS-ESI- (m/z): 410 (M + HCOO-), MS-ESI+ (m/z): 366 (M + H). TLC: toluene/EtOAc = 4:1, Rf = 0.5.
  • Early eluting products: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.20 ― 1.95 (m, 6H), 1.37 (s, 9H), 2.20 ― 2.40 (m, 1H), 2.98 (m, 2H), 3.75 (m, 1H), 3.90 (bs, 1H), 5.10 (m, 1H), 6.77 (m, 1H), 7.45 ― 7.95 (m, 5H). MS-ESI- (m/z): 410 (M + HCOO-), MS-ESI+ (m/z): 366 (M + H). TLC: toluene/EtOAc = 4:1, Rf = 0.55.
    • F. 14-O-{[1R,2R, 3S)-3-(2-tert-Butoxycarbonylamino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S,2S,3R) diasteromer thereof
  • Figure imgb0040
  • To a solution of 780 mg of thiobenzoic acid S-[(1R,2R,SR)-5-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxy-cyclopentyl] ester and (1S, 2S, 5S) diasteromer thereof from Example 7 Step E in 10 mL of acetonitrile was added 210 µL of hydrazine monohydrate and the solution obtained was stirred for 30 min at rt. After addition of 25 mL of 1 M HCl the mixture obtained was stirred for 5 min, diluted with DCM and the layers were separated. The organic layer obtained was dried and solvent was evaporated. The evaporation residue obtained was dissolved in 10 mL of acetonitrile. To the mixture obtained 279 µL of DBN and 1.13 g of pleuromutilin tosylate was added and the solution obtained was stirred for 1 h at rt. To the mixture obtained DCM was added, the phases obtained were separated and the organic phase was washed with brine, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, DCM /MeOH = 99:1) to obtain 546 mg of Example 7 Step F products in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.82 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.42 (bs, 1H), 2.92 (m, 2H), 3.02 (m, 1H), 3.23 (m, 2H), 3.43 (m, 1H), 3.79 (m, 1H), 4.55 (d, 1H, J=6Hz), 4.76 (m, 1H), 5.08 (m, 2H), 5.55 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 6.77 (t, 1H, J=6Hz). MS-ESI+ (m/z): 622 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.5.
    • G. 14-O-{[1R, 2R, 3S)-2-Hydroxy-3-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl} -mutilin and the (1S, 2S, 3R) diastereomer thereof
  • Figure imgb0041
  • 530 mg (0.85 mmol) of Example 7 Step F products was dissolved in 5 mL of DCM and 260 µL of TFA was added. After 2 d at rt the reaction mixture was charged with DCM and the organic layer obtained was extracted with saturated NaHCO3 solution. The organic layer obtained was dried and solvent was evaporated and the evaporation residue obtained wasmsubjected to chromatography (silica, DCM / MeOH / NH4OH = 95:5:0.5 - 4:1:0.1) to obtain 75 mg of Example 7 Step G products in the form of a white foam together with 384 mg of a mixed fraction containing starting material.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.66 (d, 3H, J=6Hz), 0.84 (d, 3H, J=6Hz), 1.08 (s, 3H), 1.39 (s, 3H), 2.44 (bs, 1H), 3.06 (m, 1H), 3.20 (m, 2H), 3.30 (m, 2H), 3.45 (m, 1H), 3.83 (m, 1H), 4.57 (d, 1H, J=6Hz), 4.90 (m, 1H), 5.09 (m, 2H), 5.57 (d, 1H, J=8Hz), 6.16 (dd, 1H, J=11 and 18Hz), 9.43 (t, 1H, J=4Hz). MS-ESI- (m/z): 662 (M + HCOO-), MS-ESI+ (m/z): 635 (M + NH4). TLC: DCM/MeOH = 95:5, Rf = 0.45.
    • Example 8 14-O-{[(1R, 2R, 3S)-3-(2-Amino-ethyl)-2-hydroxy cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 3R) diastereomer A. 14-O-{[(1R, 2R, 3S)-3-(2-Amino-ethyl)-2-hydroxy cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer
  • Figure imgb0042
  • To a solution of 320 mg of Example 7 Step F products in 3 mL of MeOH was added 1 mL of 28% NH4OH at rt. After 24 h a further 1 mL of 28% NH4OH was added and the mixture obtained was stirred for additional 24 h. From the mixture obtained solvent was evaporated, the evaporation residue obtained was dissolved in DCM and the mixture obtained was extracted with saturated aqueous NaHCO3 solution. The organic phase obtained was dried, the solvent was evaporated and the evaporation residue obtained was subjected to chromatography (silica, DCM/MeOH = 9:1 ― DCM/MeOH/NH4OH = 7:1:0.1) to obtain 88 mg of free base Example 8 Step A products in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.66 (d, 3H, J=6Hz), 0.84 (d, 3H, J=6Hz), 1.08 (s, 3H), 1.39 (s, 3H), 2.44 (bs, 1H), 3.07 (m, 1H), 3.28 (m, 2H), 3.45 (m, 1H), 3.81 (m, 1H), 4.58 (d, 1H, J=6Hz), 5.08 (m, 2H), 5.57 (d, 1H, J=8Hz), 6.16 (dd, 1H, J=11 and 18Hz). MS-ESI- (m/z): 566 (M + HCOO-), MS-ESI+ (m/z): 522 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.30.
    • B. 14-O-{[(1R, 2R,3S)-3-(2-Amino-ethyl)-2-hydroxy cyctopentylsutfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 3R) diastereomer
  • Figure imgb0043
  • 75 mg of the free base from Example 8 Step A was dissolved in 5 mL of DCM and 1 mL of 1 M HCl in diethylether was added. After 10 min the solvent was evaporated and the hydrochlorides of the Example 8 title compounds were obtained in a quantitative manner.
  • MS-ESI- (m/z): 556 (M + Cl-), MS-ESI+ (m/z): 522 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.30.
    • Example 9 14-O-{[(1R,2R, 5R)-2-Hydroxy-5-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof and 14-O-{[(1R, 2R, 5R)-5-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof A. 14-O- {[(1R,2R,5R)-5-(2-tert-Butoxycarbonylamino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acelyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof
  • Figure imgb0044
  • To a solution of 1.17 g of thiobenzoic acid S-[(1R, 2R, 5R)-5-(2-tert-butoxycarbonylamino-ethyl)-2-hydroxy-cyclopentyl) ester and the (1S, 2S, 5S) diastereomer thereof from Example 7 Step E in 20 mL of acetonitrile was added 316 µL of hydrazine monohydrate and the solution obtained was stirred for 30 min at rt. After addition of 25 mL of 1 M HC1 the mixture obtained was stirred for 5 min, diluted with DCM and the layers obtained were separated. The organic layer obtained was dried and the solvent was evaporated. The evaporation residue obtained was dissolved in 20 mL of acetonitrile. To the mixture obtained 420 µL of DBN and 1.53 g of pleuromutilin tosylate were added and the solution obtained was stirred for 2 h at rt. The mixture obtained was charged with DCM, the phases obtained were separated and and the organic phase obtained was washed with brine, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, DCM/MeOH = 98:2 ― 95:5) to obtain 1.21 g of Example 9 Step A products in the form of a a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.65 (d, 3H, J=6Hz), 0.84 (d, 3H, J=6Hz), 1.08 (s, 3H), 1.38 (s, 12H), 2.43 (bs, 1H), 2.56 (m, 1H), 2.90 (m, 2H), 3.34 (m, 2H), 3.46 (m, 1H), 3.92 (m, 1H), 4.56 (d, 1H, J=6Hz), 4.92 (t, 1H, J=4Hz), 5.09 (m, 2H), 5.57 (d, 1H, J=8Hz), 6.16 (dd, 1H, J=11 and 18Hz), 6.81 (m, 1H). MS-ESI+ (m/z): 622 (M + H). TLC: DCM/MeOH = 95:5, Rf = 0.45.
    • B. 14-O-{[(1R, 2R, 5R)-2-Hydroxy-5-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof and 14-O-{[(1R,2R,5R)-5-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof
  • Figure imgb0045
  • To a solution of 1.16 g (1.86 mmol) of the Example 9 Step A products from above in 10 mL of DCM was added 285 µL of TFA and the mixture obtained was stirred for 24 h at rt. Further 300 µL of TFA was added and the reaction mixture was stirred for additional 24 h at rt. To the mixture obtained DCM and saturated NaHCO3 solution was added, and the phases obtained were separated. The organic phase obtained was extracted with DCM, dried and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, DCM/MeOH = 98:2 ― DCM/MeOH/NH4OH = 9:1:0.1 ― DCM/MeOH/NH4OH = 4:1:0.2) to obtain 50 mg of 14-O-{[(1R, 2R, 5R)-2-Hydroxy-5-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof in the form of a white foam as well as 142 mg of 14-O-{[(1R, 2R, 5R)-5-(2-Aminoethyl)-2-hydroxy-cyclopentylsulfanylacetyl]-mutilin and the (1S, 2S, 5S) diastereomer thereof in the form of a white foam.
  • Acetamide: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.65 (d, 3H, J=6Hz), 0.83 (d, 3H, J=6Hz), 1.07 (s, 3H), 1.39 (s, 3H), 2.43 (bs, 1H), 2.58 (m, 1H), 3.20 (m, 2H), 3.48 (m, 3H), 3.94 (m, 1H), 4.55 (m, 1H), 4.94 (t, 1H, J = 4Hz), 5.09 (m, 2H), 5.57 (d, 1H, J=8Hz), 6.16 (dd, 1H, J=11 and 18Hz), 9.45 (m, 1H). MS-ESI- (m/z): 662 (M + HCOO-), MS-ESI+ (m/z): 635 (M + NH4). TLC: DCM/MeOH = 95:5, Rf = 0.4.
    Free base: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.66 (d, 3H, J=6Hz), 0.84 (d, 3H, J=6Hz), 1.08 (s, 3H), 1.40 (s, 3H), 2.44 (bs, 1H), 2.65 (m, 3H), AB-system (υA= 3.44, υB= 3.29, J = 28Hz), 3.48 (m, 1H), 3.95 (m, 1H), 4.59 (bs, 2H, J=6Hz), 5.08 (m, 2H), 5.57 (d, 1H, J=8Hz), 6.17 (dd, 1H, J=11 and 18Hz). MS-ESI- (m/z): 566 (M + HCOO-), MS-ESI+ (m/z): 522 (M + H). TLC: DCM/MeOH = 95:5 + 1% NH4OH, Rf = 0.35.
    • Example 10 14-O-{[(1R,2R, 5R)-5-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S) diastereomer thereof
  • Figure imgb0046
  • 100 mg of the free base from Example 9 Step B was dissolved in 5 mL of DCM and 1 mL of 1 M HCl in dietheylether was added. After 10 min the solvent was evaporated and the hydrochlorides of the Example 10 title compounds were obtained in quantitative yield.
  • MS-ESI- (m/z): 556 (M + Cl-), MS-ESI+ (m/z): 522 (M + H). TLC: DCM/MeOH = 95:5 + 1% NH4OH, Rf = 0.35.
    • Example 11 14-O-{[(1R, 2R, 5R)-5-[2-(2,2-Dimethyl-propylamino)-ethyl]-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S) diastereomer thereof A. 14-O-{[(1R,2R,5R)-5-[2-(2,2-Dimethyl-propylamino)-ethyl]-2-hydroxy-cyclopentylsulfany]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof
  • Figure imgb0047
  • 104 mg of 14-O-{[(1R, 2R, 5R)-5-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof from Example 9 Step B was dissolved in 2 mL of dry DCM containing molecular sieves. To the mixture obtained 22 mg of pivalaldehyde was added at rt and the reaction mixture obtained was stirred for 2 h. Then To the mixture obtained 55 mg of sodium triacetoxyborohydride was added and stirring was continued for another 18 h. To the mixture obtained DCM was added, the phases obtained were separated and the organic phase obtained was washed with aqueous saturated NaHCOs solution, dried and the solvent was evaporated. After chromatography of the residue (silica, DCM/MeOH = 98:2 ― 95:5 - 9:1) 58 mg of Example 11 Step A products was obtained in the form of a white foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.66 (d, 3H, J=6Hz), 0.84 (d, 3H, J=6Hz), 0.91 (s, 9H), 1.09 (s, 3H), 1.40 (s, 3H), 2.44 (bs, 1H), 3.30 ― 3.50 (m, 4H), 3.94 (m, 1H), 4.58 (d, 1H, J=6Hz), 4.97 (m, 1H), 5.10 (m, 2H), 5.58 (d, 1H, J=8Hz), 6.17 (dd, 1H, J=11 and 18Hz). MS-ESI+ (m/z): 592 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.45.
    • B. 14-O-{[(1R,2R, 5R)-5-[2-(2,2-Dimethyl-propylamino)-ethyl]-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S) diastereomer thereof
  • 47 mg of the free base from Example 11 Step A was dissolved in 3 mL of DCM and 300 µL of 2 M HCl in diethylether was added. The mixture obtained was stirred for 10 min and solvent was evaporated. 48 mg of the hydrochloride Example 11 title compounds was obtained in the form of a white solid.
    MS-ESI+ (m/z): 592 (M + H). TLC: DCM/MeOH = 9:1 + 1% NH4OH, Rf = 0.45.
    • Example 12 14-O-{[(1R, 2R, 5R)-5-Amino-2-hydroxy-cycloheptylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S), (1R, 2R, 5S), (1S, 2S, 5R) diastereomers thereof A. tert-Butyl cyclohept-4-enyl-carbamate
  • Figure imgb0048
  • A solution of 3.5 g of LAH in 180 mL of THF was heated to reflux and 90.6 mmol of cyclohept-4-enone (Louie, J. et al, J. Am. Chem. Soc. 2001, 123, 11312-11313) in 20 mL of THF were added over 45 min. After further 4 h at reflux the mixture obtained was allowed to cool to rt, stirred overnight at rt, and was charged with diluted HCl and 100 mL of EtOAc. The phases obtained were separated and the aqueous layer was extracted with EtOAc. The combined organic layers obtained were washed with saturated aqueous NaHCO3, dried over sodium sulfate and filtered. After chromatography (silica, CH/EtOAc = 2.5/1) 370 mg of cyclohept-4-enol was obtained in the form of a brown oil.
    MS-ESI+ (m/z): 130 (M + NH4). TLC: CH/EtOAc = 2:1, Rf = 0.4.
  • A solution of 360 mg of cyclohept-4-enol in 10 mL of THF was charged portionwise with 0.88 mL of N-methylmorphiline and 1062 mg of methanesulfonic anhydride. Upon completion of the reaction, the mixture obtained was distributed between EtOAc and saturated aqueous NaHCO3, and the aqueous layer obtained was extracted with EtOAc. The combined organic layers obtained were washed with 1N HCl, water and brine, dried over sodium sulfate and filtered. The filtrate obtained was concentrated to dryness to give the corresponding mesylate in the form of a yellow oil in quantitative yield.
    MS-ESI+ (m/z): 208 (M + NH4), 398 (2M + NH4). TLC: CH/EtOAc = 2:1, Rf = 0.5.
  • 3.2 mmol of the mesylate from above was heated to 60 °C overnight with 840 mg of sodium azide in 10 mL of DMF. The reaction mixture obtained was charged with water and extracted twice with diethylether. The combined organic layers obtained containing the crude azide were concentrated and used for the next Step.
    MS-ESI- (m/z): 136 (M - H). TLC: CH/EtOAc = 3:1, Rf = 0.9.
  • To the crude azide from above in 10 mL of THF and 0.5 mL of water was added 1 g of triphenylphosphine. The reaction mixture obtained was stirred at 80 °C for 4 h, cooled to rt and charged with 0.8 mL of Boc anhydride in 3 mL of THF. Upon completion of the reaction, 1 M HCl was added and the mixture obtained was extracted twice with EtOAc. The combined organic layers obtained were washed with water and brine, dried over sodium sulfate and filtered. After chromatography (silica, CH) the Example 12 Step A products was obtained in the form of a yellow oil.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, chatracteristic signals): 1.35 (s, 9H), 3.48 (m, 1H), 5.70 (m, 2H), 6.88 (d, 1H, J=8Hz). TLC: CH, Rf = 0.5.
    • B. 14-O-{[(1R,2R,5R)-5-tert-Butoxycarbonylamino-2-hydroxy-cycloheptylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S), (1R, 2R, 5S), (1S, 2S, 5R) diastereomers thereof
  • Figure imgb0049
  • To a solution of 3.2 mmol of Example 12 Step A product in 20 mL of DCM was added 980 mg of mCPBA (70%) and 350 mg of NaHCO3. The resulting suspension was stirred at rt overnight, then charged with 20% sodium thiosulfate and extracted with EtOAc. The combined organic layers obtained were washed with saturated aqueous NaHCO3, water and brine, dried over sodium sulfate and filtered. The filtrate obtained was concentrated and dried to obtain crude epoxide in the form of a yellowish solid in quantitiative yield.
    TLC: CH/EtOAc = 1:1, Rf = 0.7.
  • 540 mg of the epoxide obtained above was dissolved in 10 mL of toluene, charged with 0.6 mL of thiobenzoic acid and 60 mg of Bu4NCl and the mixture obtained was stirred at 50 °C overnight. Upon completion of the reaction, the mixture obtained was distributed between EtOAc and saturated aqueous NaHCO3, and the aqueous layer obtained was extracted with EtOAc. The combined organic layers obtained were washed with water and brine, dried over sodium sulfate and filtered. The filtrate obtained was concentrated to dryness to give a diastereomeric mixture of thiobenzoic acid S-((1R, 2R, 5R)-5-tert-butoxycarbonylamino-2-hydroxy-cycloheptyl) ester and the (1R, 2R, 5S), (1S, 2S, 5S), (1S, 2S, 5R) diastereomers in the form of a a brown oil.
    TLC: CH/EtOAc = 1:1, Rf = 0.7.
  • To a solution of the mixture of thiobenzoic esters obtained in 10 mL of DCM was added 60 mg of DTT and 0.2 mL of hydrazine monohydrate and the mixture obtained was stirred for 6 h at rt. Upon completion of the reaction, 1 M phosphoric acid was added and the mixture obtained was extracted twice with EtOAc. The combined organic layers obtained were washed with 1% aqueous NaCl solution, dried over sodium sulfate, filtered, concentrated and immediately used for the next reaction.
    To the crude thiol obtained above was added 1.2 g of pleuromutilin tosylate in 6 mL of MTBE, 75 mg benzyltributylammonium chloride and 2.5 mL of 1 M NaOH and the resulting biphasic mixture was stirred over the weekend at rt. The mixture obtained was extracted with EtOAc and water and the organic layer obtained was dried over sodium sulfate, filtered and concentrated. After chromatography (silica, CH/EtOAc = 5/1) 610 mg of a mixture of the Example 12 Step B products was obtained in the form of a colorless foam.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 0.62 (d, 3H, J=6Hz), 0.82 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.36 (s, 12H), 2.40 (bs, 1H), 3.56 (m, 1H), 4.56 (d, 1H, J=6Hz), 4.84 (d, 1H, J=6Hz), 5.05 (m, 2H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 Hz and 17Hz), 7.6 (m, 1H). MS-ESI- (m/z): 667 (M + HCOO-).
    • C. 14-O-{[(1R, 2R, 5R)-5-Amino-2-hydroxy-cycloheptylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2S, 5S), (1R, 2R, 5S), (1S, 2S, 5R) diastereomers thereof
  • Figure imgb0050
  • A solution of 600 mg of Example 12 Step B products in 10 mL of DCM was treated with 1 mL of TFA for 7 h at rt. Then 1 M NaOH was added and the resulting mixture was extracted twice with EtOAc. The combined organic layers obtained were washed with saturated aqueous NaHCO3 solution, water and brine, dried over sodium sulfate, filtered and concentrated under reduced pressure. After chromatography (silica, EtOAc / MeOH/ NH40H = 75/15/1) 44 mg of a mixture of the Example 12 Step C products was obtained in the form of a colorless foam.
  • 1H NMR (400MHz, DMSO-d6, δ, ppm, characteristic signals): 0.62 (d, 3H, J=6Hz). 0.81 (d, 3H, J=7Hz), 1.05 (s, 3H), 1.36 (s, 3H), 2.40 (bs, 1H), 2.82 (m, 1H), 2.99 (m, 1H), 3.64 (m, 1H), 4.50 (m, 1H), 5.05 (m, 2H), 5.54 (d, 1H, J=8Hz), 6.13 (m, 1H). MS-ESI+ (m/z): 522 (M + H), MS-ESI- (m/z): 520 (M - H). TLC: EtOAc/MeOH = 9:1 + 1% NH4OH, Rf = 0.3.
    • D. 14-O-{[(1R, 2R, 5R)-5-Amino-2-hvdroxy-cycloheptvlsulfanyl]-acetyl}-mutilin hydrochloride hydrochloride and the (1S, 2S, 5S), (1R, 2R, 5S), (1S, 2S, 5R) diastereomers thereof
  • Figure imgb0051
  • To a solution of 40 mg of Example 12 Step C products in 1 mL of dioxane was added 0.12 mL of 1 M HCl. After stirring at rt for 10 minutes the solution obtained was lyophilized to obtain Example 12 Step D products in the form of a colorless foam in quantitative yield. MS-ESI+ (m/z): 522 (M +). TLC: EtOAc/MeOH = 9:1 + 1% NH4OH, Rf = 0.3.
    • Example 13 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Amino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof A. (1S, 5R, 6S)-Bicyclo[3.2.0]hept-2-en-6-yl-(2,4-dimethoxy-benzyl)-amine and the (1R, 5S, 6R) diasteromer thereof
  • Figure imgb0052
  • To a solution of 10 g of cis bicyclo[3.2.0]hept-2-en-6-one in 100 mL of dry DCM was added 185 mmol 2,4-dimethoxybenzylamine, 8 mL of acetic acid and 27 g of sodium triacetoxyborohydride at rt and the resulting reaction mixture was stirred overnight. To the mixture obtained EtOAc was added, washed with 1 N NaOH and the aqueous phase was extracted with EtOAc. The combined organic layers obtained were washed with water and brine, dried and evaporated. After chromatography (silica, EtOAc / MeOH = 5/1) 21.64 g of Example 13 Step A products was obtained in the form of a yellow oil.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.70 (m, 1H), 2.10 ― 2.70 (m, 4H), 2.95 (m, 2H), 3.40 (m, 2H), 3.73, 3.76 (2s, 6H), 5.77 (m, 2H), 6.40 ― 7.20 (m, 3H). MS-ESI+ (m/z): 260 (M + H). TLC: EtOAc/MeOH = 1:1, Rf = 0.5.
    • B. tert-Butyl (1S, 5R, 6S)-Bicyclo[3.2.0]hept-2-en-6-yl-(2,4-dimethoxy-benzyl)-carbamate and the 1R, 5S, 6R) diasteromer thereof
  • Figure imgb0053
  • To a solution of 21.64 g of Example 13 Step A products in 150 mL of DCM was added 22 mL of Boc-anhydride and 25.3 mL of TEA and the resulting reaction mixture was stirred at rt overnight. From the mixture obtained solvent was evaporated and the evaporation residue obtained was charged with EtOAc. The mixture obtained was washed with 1 M HC1, the organic layer obtained was dried over sodium sulfate and the solvent was evaporated to obtain 30.46 g of Example 13 Step B products in the form of a yellow solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.36 (s, 9H), 1.58 (m, 1H), 2.20 ― 2.45 (m, 3H), 2.90 (m, 1H), 3.07 (m, 1H), 3.72, 3.75 (2s, 6H), 4.10 (d, 1H J=17Hz), 4.42 (m, 1H), 4.53 (d, 1H, J=17Hz), 5.73 (m, 1H), 5.83 (m, 1H), 6.40 ― 6.84 (m, 3H). MS-ESI+ (m/z): 360 (M + H). TLC: CH/EtOAc = 3:1, Rf = 0.65.
    • C. tert-Butyl (R)-(2,4-Dimethoxy-benzyl)-(1R,3S,6R, 7S)-3-oxa-tricyclo[4.2.0*2,4*]oct-7-yl-carbamate and the (1S, 3R, 6S, 7R) diasteromer thereof
  • Figure imgb0054
  • 10.4 g of mCPBA (70%) was added at 0 °C to a solution of 15.2 g of Example 13 Step B products in 100 ml of DCM. To the mixture obtained 5.3 g ofNaHCO3 was added and the mixture obtained was stirred at rt for 2 days. The mixture obtained was charged with 20% sodium thiosulfate, stirred for 10 min and extracted with EtOAc. The combined organic layers obtained were washed with saturated NaHCO3 solution, water and brine, dried over sodium
    sulfate and solvent was evaporated. The crude evaporation residue obtained was dried under HV to yield 16.9 g of Example 13 Step C products in the form of a brown oil.
    MS-ESI+ (m/z): 376 (M + H). TLC: CH/EtOAc = 2:1, Rf = 0.5.
    • D. Thiobenzoic acid S-{[(1R, 2s, 3S, 5R, 6S)-6-[tert-butoxycarbonyl-(2,4-dimethoxybenzyl)-amino]-2-hydroxy-bicyclo[3.2.0]hept-3-yl} ester and the (1S, 2R, 3R, 5S, 6R) diasteromer thereof
  • Figure imgb0055
  • 10.5 mL of thiobenzoic acid was added dropwise to a solution of 16.9 g of epoxide from Example 13 Step C in 100 mL of dry toluene containing 1 g of Bu4NCl. The reaction mixture obtained was stirred for 20 h at rt and for 20 h at 55 °C. To the mixture obtained saturated NaHCO3 solution was added and after 20 min the reaction mixture was charged with EtOAc. The phases obtained were separated and and the organic phase was washed 3 times with saturated aquepus NaHCO3 solution, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 4: 1) to obtain 6.49 g of Example 13 Step D products in the form of an oil.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.37 (s, 9H), 3.72, 3.77 (2s, 6H), 3.86 (m, 2H), 4.21 (d, 1H J=l7Hz), 4.38 (d, 1H, J=l7Hz), 5.17 (d, 1H, J=5Hz), 6.40 - 6.95 (m, 3H), 7.50 - 7.95 (m, 5H). MS-ESI- (m/z): 558 (M + HCOO-), MS-ESI+ (m/z): 514 (M + H). TLC: CH/EtOAc = 2:1, Rf = 0.35.
    • E. 14-O- [(1R, 2S, 3S, 5R, 6S)- 6-[tert-butoxycarbonyl-(2,4-dimethoxy-benzyl)-amino]-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetvl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • To a solution of 6.49 g of Step D products from above in 20 mL of DCM was added at 0 °C 649 mg of DTT and 0.9 mL of hydrazine monohydrate and the solution obtained was stirred for 2 h at rt. After addition of 1 M H3PO4 the mixture obtained was diluted with DCM and the layers were separated. The organic layer obtained was washed 3 times with 1 M H3PO4, once with 1% NaCl solution, dried over sodium sulfate and the solvent was evaporated. MS-ESI+ (m/z): 410 (M + H). MS-ESI- (m/z): 408 (M - H). TLC: CH/EtOAc = 1:1, Rf = 0.4.
  • The evaporation residue obtained was dissolved in 50 mL of dry acetonitrile and 1.7 mL of DBN and 6.06 g of pleuromutilin tosylate were added. The resulting mixture was stirred for 2 days at rt, charged with EtOAc, the phases obtained were separated and the organic phase obtained was washed with brine, dried and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 2:1) to yield 4.98 g of Example 13 Step E products in the form of a white foam.
  • 1H-NMR (400 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.65 (d, 3H, J=8Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.41 (bs, 1H), 2.93 (m, 1H), 3.05 (m, 1H), 3.44 (t, 1H, J=4Hz), 3.70 (t, 2H, J=5Hz), 3.80 (m, 1H), 4.15 (d, 1H, J=l2Hz), 4.37 (d, 1H, J=l2Hz), 4.51 (d, 1H, J=7Hz), 5.07 (m, 2H), 5.56 (d, 1H, J=8Hz), 6.15 (m, 1H), 6.45 - 6.90 (m, 3H). MS-ESI+ (m/z): 770 (M + H). TLC: CH/EtOAc = 1:2, Rf = 0.55.
    • F. 14-0-{[(1R, 2S, 3S, 5R, 6S)- 6-Amino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0056
  • To a solution of 4.98 g of Example 13 Step E products from above in 20 mL of DCM was added at 0 °C 2.5 mL of TFA, 0.3 mL of thioanisole and 0.13 mL of methanesulfonic acid and the solution obtained was stirred for 6 h at rt. The reaction mixture obtained was added dropwise under stirring to cold DIPE and the resulting precipitate was filtered and dried. The solid was taken up in EtOAc, the pH adjusted to 12 with 5 M NaOH and the aqueous layer obtained was extracted with EtOAc. The combined organic layers obtained were dried over sodium sulfate and the solvent was evaporated to obtain 1.98 g of free base Example 13 Step F products in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.36 (s, 3H), 2.41 (bs, 1H), 2.77 (m, 1H), 3.04 (m, 1H), 3.28 (m, 1H), 3.41 (m, 3H), 3.70 (m, 1H), 4.54 (bs, 1H), 5.06 (m, 2H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz). MS-ESI+ (m/z): 520 (M + H). TLC: EtOAc/MeOH = 10:1 + 1% NH4OH, Rf = 0.4.
    • G. 14-0-{[(1R, 2S, 3S, 5R, 6S)- 6-Amino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0057
  • To a solution of 300 mg of free base Example 13 Step F products in 1 mL of dioxane was added 0.9 mL of 1 M HCI. After stirring at rt for 1 minutes the solution obtained was lyophilized to obtain 301 mg of the Example 13 title compounds in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.41 (bs, 1H), 2.92 (bm, 2H), 3.47 (m, 3H), 3.74 (m, 1H), 4.56 (bd, 1H), 5.10 (m, 3H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz). MS-ESI- (m/z): 554 (M + C1-), 564 (M + HCOO-), MS-ESI+ (m/z): 520 (M + H).
    • Example 14 14-0-{[(1R, 2S, 3S, 5R, 6S)-6-Formylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0058
  • To a solution of 0.1 mL acetic formic anhydride in DCM was added 71 mg of DMAP and 300 mg of free base from Example 13 Step F. After stirring at rt for 2 days the solution obtained was charged with EtOAc, the mixture obtained was washed with brine, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 1:10) to obtain 100 mg of the Example 14 title compounds in the form of a colorless solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.05 (s, 3H), 1.37 (s, 3H), 2.41 (bs, 1H), 2.80 - 3.10 (m, 2H), 3.41 (m, 3H), 3.73 (m, 1H), 4.20 (m, 1H), 4.54 (d, 1H, J=6Hz), 5.07 (m, 3H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 7.89 (s, 1H), 8.04 (d, 1H, J=7Hz). MS-ESI- (m/z): 592 (M + HCOO-), MS-ESI+ (m/z): 548 (M + H). TLC: toluene/EtOAc = 1:10, Rf = 0.2.
    • Example 15 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-(2-Amino-acetylamino)-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof A. 14-0-{[(1R, 2S, 3S, 5R, 6S)-6-(2-tert-Butoxycarbonylamino-acetylamino)-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0059
  • To a solution of 102 mg of Boc-glycine in DCM was added 78 mg of HOBT, 71 mg of DMAP, 111 mg of EDC and the resulting mixture was stirred for 30 min at rt. To the mixture obtained the 300 mg of free base from Example 13 Step F was added. After stirring at rt for 2 days the solution obtained was charged with EtOAc, the phases obtained were separated and the organic phase obtained was washed with brine, dried and the solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, toluene/EtOAc = 1:20) to obtain 196 mg of the Example 15 Step A products in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=5Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.41 (bs, 1H), 2.99 (m, 1H), 3.04 (m, 1H), 3.45 (m, 5H), 3.74 (m, 1H), 4.18 (m, 1H), 4.54 (d, 1H, J=6Hz), 5.07 (m, 3H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=10 and 17Hz), 6.92 (m, 1H), 7.76 (d, 1H, J=8Hz). MS-ESI- (m/z): 721 (M + HCOO-), MS-ESI+ (m/z): 677 (M + H). TLC: toluene/EtOAc = 1:10, Rf = 0.25.
    • B. 14-0-{[(1R, 2S, 3S, 5R, 6S)-6-(2-Amino-acetylainino)-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0060
  • To a solution of 196 mg of the Example 15 Step A products from above in 3 mL of DCM was added 0.3 mL of TFA and the mixture obtained was stirred for 6 h at rt. To the mixture obtained EtOAc and 1 M NaOH were added, the phases obtained were separated and the organic phase obtained was extracted with EtOAc and the combined organic layers obtained were washed with water and brine, dried and the solvent was evaporated. After chromatography of the residue (silica, EtOAc/MeOH/NH4OH = 90:3:1) 68 mg of the free base Example 15 Step B products was obtained in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.37 (s, 3H), 2.41 (bs, 1H), 2.92 (m, 1H), 3.05 (m, 3H), 3.40 (m, 3H), 3.76 (m, 1H), 4.22 (m, 1H), 4.54 (d, 1H, J=5Hz), 5.06 (m, 3H), 5.56 (d, 1H, J=8Hz), 6.15 (dd, 1H, J=11 and 18Hz), 7.95 (d, 1H, J=8Hz). MS-ESI- (m/z): 621 (M + HCOO-), MS-ESI+ (m/z): 577 (M + H). TLC: EtOAc/MeOH = 10:1 + 1% NH4OH, Rf = 0.2.
    • C. 14-O-H{[(1R, 2S, 3S, 5R, 6S)-6-(2-Amino-acetylamino)-2-hydroxy-bicyclor[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin hydrochlorides and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0061
  • To a solution of 68 mg of free base Step B products in dioxane was added 0.36 mL of 1 M HC1. After stirring at rt for 10 minutes the solution obtained was lyophilized to obtain 64 mg of the Example 15 title compounds in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.36 (s, 3H), 2.41 (bs, 1H), 2.95 (m, 2H), 3.75 (m, 1H), 4.20 (m, 4H), 5.06 (m, 2H), 5.56 (d, 1H, J=8Hz), 6.14 (dd, 1H, J=11 and 18Hz), 8.10 (m 3H), 8.50 (m, 1H). MS-ESI- (m/z): 611 (M + Cl-), 621 (M + HCOO-), MS-ESI+ (m/z): 577 (M + H).
    • Example 16 14-0-{[(1R, 2S, 3S, 5R, 6S)-6-Cyclopropylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof A. (1S, 5R, 6S)-Bicyclo[3.2.0]hept-2-en-6-yl-cyclopropyl-amine and the (1R, 5S, 6R) diasteromer thereof
  • Figure imgb0062
  • To a solution of 5 g of cis bicyclo[3.2.0]hept-2-en-6-one in 50 mL of dry DCM was added 3.22 mL of cyclopropylamine, 2.65 mL of acetic acid and 13.7 g of sodium triacetoxyborohydride at rt. After 18 h the solvent was evaporated, EtOAc was added to the evaporation residue obtained, the mixture obtained was washed with 1 N NaOH and the aqueous phase obtained was extracted with EtOAc. The combined organic layers obtained were washed with water and brine, dried and solvent was evaporated. After chromatography (silica, EtOAc / MeOH = 5/1) 4.02 g of Example 16 Step A products was obtained in the form of a colorless oil.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.05 - 0.40 (m, 4H), 1.29 (m, 1H), 1.91 (m, 1H), 2.10 - 2.70 (m, 3H), 2.97 (m, 2H), 3.46 (m, 1H), 5.76 (m, 2H). MS-ESI+ (m/z): 150 (M + H). TLC: CH/EtOAc = 4:1, Rf = 0.35.
    • B. tert-Butyl (1S, 5R, 6S)-Bicyc1o[3.2.0]hept-2-en-6-yl - cycopropyl-carbamate and the (1R, 5S, 6R) diasteromer thereof
  • Figure imgb0063
  • To a solution of 4.02 mg of Example 16 Step A products in 100 mL of DCM was added 6.2 g of Boc-anhydride and 7.5 mL of TEA and the resulting reaction mixture was stirred at rt overnight. The mixture obtained was evaporated and the evaporation residue obtained was charged with EtOAc and the mixture obtained was washed with 1 M HCI. The organic layer obtained was dried over sodium sulfate and solvent was evaporated to obtain 7.14 g of Example 16 Step B products in the form of a colorless solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.30 - 0.80 (m, 4H), 1.37 (s, 9H), 2.00 - 2.60 (m, 5H), 2.90 (m, 1H), 3.07 (m, 1H), 4.14 (q, 1H, J=9Hz), 5.80 (m, 2H). TLC: CH/EtOAc = 4:1, Rf = 0.75.
    • C. tert-Butyl)-Cycloproply-(1R, 3S, 6R, 7S)-3-oxa-tricyclo[4.2.0*2,4*]oct-7-yl-carbamate and the (1S, 3R, 6S, 7R) diasteromer thereof
  • Figure imgb0064
  • 7.05 g of mCPBA (70%) was added at 0 °C to a solution of 7.14 g of Example 16 Step B products in 100 ml of DCM. To the mixture obtained 3.6 g of NaHCO3 was added and the mixture obtained was stirred at rt for 3 days. The mixture obtained was charged with 20% sodium thiosulfate, stirred for 20 min and extracted with EtOAc. The combined organic layers obtained were washed with saturated aqueous NaHCO3 solution, water and brine, dried over sodium sulfate and solvent was evaporated. The crude evaporation residue obtained was dried under HV to obtain 5.73 g of Example 16 Step C products in the form of a colorless solid.
    MS-ESI+ (m/z): 266 (M + H). TLC: CH/EtOAc = 2:1, Rf = 0.45.
    • D. Thiobenzoic acid S-{[(1R, 2S, 3S, 5R, 6S)-6-(tert-butoxycarbonyl-cyclopropyl-amino)-2-hydroxy-bicyclo[3.2.0]hept-3-yl} ester and the (1S, 2R, 3R, 5S, 6R) diasteromer thereof
  • Figure imgb0065
  • 20.1 mL of thiobenzoic acid was added dropwise to a solution of 5.73 g of epoxide from Example 16 Step C in 50 mL of dry toluene containing 478 mg of Bu4NCl. The reaction mixture was stirred for 3 days at rt. To the mixture obtained saturated aqueous NaHCO3 solution was added and after 15 min the mixture obtained was charged with EtOAc, the phases obtained were separated and the organic phase was washed 3 times with saturated NaHCO3 solution, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 5:1) to obtain 5.20 g of Example 16 Step D products.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.42 (m, 2H), 0.67 (m, 2H), 1.36 (s, 9H), 1.72 (m, 1H), 2.25 - 2.50 (m, 5H), 3.10 (m, 1H), 3.80 (m, 2H), 3.97 (m, 1H), 5.71 (d, 1H, J=5Hz), 7.50 -7.95 (m, 5H). MS-ESI- (m/z): 448 (M + HCOO-). TLC: CH/EtOAc = 3:1, Rf = 0.25.
    • E. 14-O-{[(1R, 2S, 3S, 5R, 6S)- 6-[tert-butoxycarbonyl-cyclopropyl-amino]-2-hydroxy bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0066
  • To a solution of 5.20 g of Example 16 Step D products in 50 mL of DCM was added at 0 °C 520 mg of DTT and 0.94 mL of hydrazine monohydrate and the solution obtained was stirred for 6 h at rt. After addition of 1 M H3PO4 the mixture obtained was diluted with DCM and the layers obtained were separated. The organic layer obtained was washed 3 times with 1 M H3PO4, once with 1% NaCl solution, dried over sodium sulfate and solvent was evaporated.
    MS-ESI+ (m/z): 300 (M + H). TLC: CH/EtOAc = 3:1, Rf = 0.2.
  • The residue was dissolved in 50 mL of dry acetonitrile and 1.7 mL of DBN and 6.20 g of pleuromutilin tosylate were added. The resulting mixture was stirred for 2 days at rt, charged with EtOAc and washed with brine. The aqueous phase was extracted with EtOAc and the combined organic layers were washed twice with brine, dried and evaporated. The residue was subjected to chromatography (silica, CH/EtOAc = 1:1) to yield 4.60 g of Example 16 Step E products as white foam.
  • 1H-NMR (400 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.36 (m, 1H), 0.46 (m, 1H), 0.56-0.72 (m, 5H), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.37 (s, 12H), 2.41 (bs, 1H), 2.97 (m, 1H), 3.08 (m, 1H), 3.43 (m, 1H), 3.70 - 3.80 (m, 2H), 4.50 (d, 1H, J=6Hz), 4.99 (bs, 1H), 5.06 (m, 2H), 5.56 (d, 1H, J=8Hz), 6.14 (m, 1H). MS-ESI- (m/z): 704 (M + HCOO-), MS-ESI+ (m/z): 660 (M + H). TLC: CH/EtOAc = 1:2, Rf = 0.5.
    • F. 14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Cyclopropylamino-2-hydroxy-bicvclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0067
  • To a solution of 4.60 g of Example 16 Step E products from above in 30 mL of DCM was added at 0 °C 6.9 mL of TFA. The resulting mixture was stirred for 6 h at rt, charged with EtOAc and saturated aqueous NaHCO3 solution. The aqueous phase obtained was extracted with EtOAc and the combined organic layers obtained were washed twice with water and brine, dried and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, EtOAc/MeOH/NH4OH = 100:5:1) to yield 3.00 g of Example 16 Step F products in the form of a colorless foam.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.10 - 0.35 (m, 4H), 0.64 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.36 (s, 3H), 2.41 (bs, 1H), 2.84 (m, 1H), 3.02 (m, 1H), 3.20 (m, 1H), 3.58 (m, 1H), 4.53 (d, 1H, J=6Hz), 5.04 (m, 3H), 5.56 (d, 1H, J=8Hz), 6.15 (m, 1H). MS-ESI+ (m/z): 560 (M + H). TLC: EtOAc/MeOH = 10:1 + 1%NH4OH, Rf=0.6.
    • G. 14-O-1[(1R, 2S, 3S, 5R, 6S)-6-Cyclopropylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin hydrochloride and the (1S, 2R, 3R, 5S, 6R) diastereomers thereof
  • Figure imgb0068
  • To a solution of 3.01 g of free base Example 16 Step F products in 10 mL of dioxane was added 8.06 mL of 1 M HCl. After stirring at rt for 10 minutes the solution obtained was lyophilized to obtain the Example 16 title compounds in the form of a colorless foam in quantitative yield.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.64 (m, 5H), 0.83 (m, 5H), 1.06 (s, 3H), 1.37 (s, 3H), 2.42 (bs, 1H), 2.94 (m, 2H), 3.73 (m, 2H), 4.57 (d, 1H, J=5Hz), 5.10 (m, 3H), 5.56 (d, 1H, J=7Hz), 6.15 (dd, 1H, J=11 and 18Hz). MS-ESI- (m/z): 594 (M + Cl-), 604 (M + HCOO-), MS-ESI+ (m/z): 560 (M + H).
    • Example 17 14-O-{[4-Acetylamino-1-hydroxy-octahydropentalen-2-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof And 14-O-{[4-Acetylamino-2-hydroxy-octahydropentalen-1-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof A. N-((3aR, 6aR)-4-lodo-octahydropentalen-1-yl)-acetamide and the (3aS, 6aS) diastereomer thereof
  • Figure imgb0069
  • To a solution of 10.33 g of iodine in 200 mL of acetonitrile was added at 0 °C 10 mL of cis,cis-1,5-cyclooctadiene and the resulting solution was stirred at rt overnight (Uemura, S. et al, J. Org. Chem. 1983, 48, 270-273). From the mixture obtained solvent was evaporated and the evaporation residue obtained was aken up in EtOAc. The mixture obtained was washed with 20% thiosulfate solution and brine, dried over sodium sulfate and solvent was evaporated. After chromatography (silica, EtOAC/MeOH = 5:1) the Example 17 Step A products were obtained in the form of a yellow solid.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 1.20 - 2.20 (m, 8H), 1.80 (s, 3H), 3.60 - 4.50 (m, 1H), 7.85 (m, 1H). TLC: CH/EtOAc = 1:3, Rf = 0.25.
    • B. (3aS, 6aR)-N-(1,2,3,3a,6,6a)-Hexahydro-pentalen-1-yl)-acetamide and the (3aS, 6aS) diastereomer thereof
  • Figure imgb0070
  • To a solution of 5.6 g of Example 17 Step A products in 50 ml of THF was added 2.14 g of potassium tert-butoxide and the resulting mixture was stirred at rt for 3 days. The mixture obtained was charged with EtOAc and washed with brine/water, dried over sodium sulfate and solvent was evaporated to obtain 4.3 g of crude Example 17 Step B products in the form of a yellow oil.
  • 1H NMR (200MHz, DMSO-d6, δ, ppm, characteristic signals): 1.00 - 2.30 (m, 6H), 1.80 (s, 3H), 2.70 (m, 1H), 2.96 (m, 1H), 3.50 (m, 1H), 5.30 (m, 2H), 7.50, 7.90 (2d, 1H). TLC: CH/EtOAc = 1:3, Rf = 0.3.
    • C. Thiobenzoic acid S-(Acetylamino-1-hydroxy-octahydropentalen-2-yl) ester and the diastereomers thereof and Thiobenzoic acid S-(Acetylamino-2-hydroxy-octahydropentalen-1-yl) ester and the diastereomers thereof
  • Figure imgb0071
  • To a solution of 2 g of Example 17 Step B products in 50 ml of DCM was added 2.98 g of mCPBA (70%) and the resulting mixture was stirred at rt overnight. The mixture obtained was diluted with DCM, washed with 20% sodium thiosulfate solution and brine, dried over sodium sulfate and solvent was evaporated to obtain the crude epoxides as a residue. That residue was taken up in dry toluene and 2.85 mL of thiobenzoic acid and 107 mg of Bu4NCl were added and the resulting mixture was stirred at rt overnight. Upon completion of the reaction, the mixture obtained was washed with aqueous saturated NaHCO3 solution, and the aqueous layer obtained was extracted with EtOAc. The combined organic layers obtained were dried over sodium sulfate and filtered. After chromatography (silica, EtOAC/MeOH = 20:1) 372 mg of the early eluting and 400 mg of the late eluting Example 17 Step C products were obtained in the form of solids.
  • Early eluting: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.20 - 2.00 (m, 8H), 1.80 (s, 3H), 2.67 (m, 1H), 4.27 (m, 2H), 5.29 (d, 1H, J=4Hz), 7.40- 8.10 (m, 6H). TLC: EtOAc, Rf = 0.25.
  • Late eluting: 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 1.20 - 2.10 (m, 8H), 1.80 (s, 3H), 350 - 4.00 (m, 2H), 5.06 (s, 1H), 7.40 - 8.05 (m, 6H). TLC: EtOAc, Rf = 0.15.
    • D. 14-O-{[4-Acetylamino-1-hvdroxy-octahydropentalen-2-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof and 14-O-{[4-Acetylamino-2-hydroxy-octahydropentalen-1-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof
  • Figure imgb0072
  • To a solution of 372 mg of Example 17 Step C early eluting products in 3 mL of DCM was added at 0 °C 20 mg of DTT and 85 µL of hydrazine monohydrate and the solution obtained was stirred for 6 h at rt. To the mixture obtained further 20 mg of DTT and 85 µL of hydrazine monohydrate were added and the mixture obtained was stirred for additional 2 h. After addition of 1 M H3PO4 the mixture obtained was diluted with DCM and the layers were separated. The organic layer obtained was washed with brine, dried over magnesium sulfate and solvent was evaporated. To the evaporation residue obtained was added 3 mL of MTBE, 1.5 mL of 1 M NaOH and 390 mg of pleuromutilin tosylate. The resulting mixture was stirred at rt overnight and diluted with EtOAc. The organic phase was separated, washed with 1 M NaOH, 0.1 M H3PO4, saturated NaHCO3 solution and brine, dried over magnesium sulfate and solvent was evaporated. The evaporation residue obtained was subjected to chromatography (silica, CH/EtOAc = 3:1 - EtOAc/MeOH/NH4OH = 500:25:1) to yield 67 mg of the Example 17 title compounds in the form of a white solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=6Hz), 0.83 (d, 3H, J=7Hz), 1.06 (s, 3H), 1.36 (s, 3H), 1.77 (s, 3H), 2.41 (bs, 1H), 3.30 - 3.60 (m, 3H), 4.00 (m, 1H), 4.55 (d, 1H, J=6Hz), 4.85 - 5.15 (m, 3H), 5.55 (d, 1H, J=8Hz), 6.16 (dd, 1H, J=11 and 18Hz), 7.65, 7.90 (bd, 1H). MS-ESI+ (m/z): 598 (M + Na). TLC: EtOAc, Rf = 0.3.
  • 400 mg of Step C late eluting products from Example 17 Step C was treated according to the method described for the early eluting Example 17 Step C products to obtain 35 mg of the Example 17 title compounds in the form of a white solid.
  • 1H-NMR (200 MHz, DMSO-d6, δ, ppm, characteristic signals): 0.63 (d, 3H, J=5Hz), 0.83 (d, 3H, J=6Hz), 1.06 (s, 3H), 1.36 (s, 3H), 1.77 (s, 3H), 2.41 (bs, 1H), 3.30 - 3.60 (m, 3H), 4.55 (d, 1H, J=6Hz), 4.85 - 5.17 (m, 3H), 5.55 (d, 1H, J=7Hz), 6.16 (dd, 1H, J=12 and 18Hz), 7.65, 7.90 (bd, 1H). MS-ESI+ (m/z): 598 (M + Na). TLC: EtOAc, Rf = 0.45.

Claims (11)

  1. A compound selected from the group of N-unsubstituted, or N-alkylated, or N-acylated
    14-O-[(amino(C0-4)alkyl-hydroxy-cycloalkyl- and bicycloalkylsulfanyl)-acetyl]-mutilins which are
    14-O-[(amino(C0-4)alkyl-hydroxy-cyclobutytsulfanyl)-acetyl]-mutilins,
    14-O-[(amino(C0-4)alkyl-hydroxy-cyclopentylsulfanyl)-acetyl]-mutilins,
    14-O-[(amino(C0-4)alkyl-hydroxy-cycloheptylsulfanyl)-acetyl]-mutilins,
    14-O-[(amino(C0-4)alkyl-hydroxy-cyclooctansulfanyl)-acetyl]-mutilins, and
    14-O-[(amino(C0-4)alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilins.
  2. A compound according to claim 1 which is an N-unsubstituted, or N-alkylated, or N-acylated 14-O-[amino(C-4)alkyl-hydroxy-cydopentylsulfanyl)-acetyl]-mutilin or a 14-O-[amino(C0-4)alkyl-hydroxy-bicycloalkylsulfanyl)-acetyl]-mutilin wherein one ring of the bicyclyic alkyl is a cyclopentyl ring.
  3. A compound according to claim 1 of formula (I)
    Figure imgb0073
     or of formula (II)
    Figure imgb0074
     wherein
    m is 0, 1, 2, 3 or 4;
    n is 0, 1, 3 or 4;
    o is0, 1, 2, 3 or 4;
    p is 0, 1, 2, 3 or 4;
    R is ethyl or vinyl;
    R1 is hydrogen or (C1-6)alkyl,
    R2 is hydrogen or
    - (C3-6)cycloalkyl, or
    - unsubstituted (C1-6)alkyl, or
    - (C1-6)alkyl substituted by one or more of
    - hydroxy; preferably one or two,
    - methoxy,
    - halogen,
    - (C3-6)cycloalkyl, or
    R1 and R2 together with the nitrogen atom to which they are attached form a 5 to 7 membered heterocyclic ring containing at least 1 nitrogen atom or 1 nitrogen and 1 additional heteroatom e. g. selected from N or O, or R2 is a group of formula
    Figure imgb0075
     wherein
    R3 is hydrogen, straight chain or branched (C1-8)alkyl or (C3-8)cycloalkyl, or
    R3 is that part of an natural amino acid in D or in L form which remains if the carboxylic acid group is split off, or
    R3 is that part of a non natural amino acid in D or in L form which remains if the carboxylic acid group is split off.
  4. A compound according to claim 1 of formula (III)
    Figure imgb0076
     or of formula (IV)
    Figure imgb0077
     wherein m, n, o, p, R1 and R2 are as defined in claim 1.
  5. A compound according to claim 1 of formula (V)
    Figure imgb0078
     or of formula (VI)
    Figure imgb0079
     or of formula (VII)
    Figure imgb0080
     or of formula (VIII)
    Figure imgb0081
     wherein m, R1 and R2 are as defined in claim 1.
  6. A compound according to any one of claims 1 to 5, selected from the group consisting of
    14-O-{[(1R, 2R, 4R)-4-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof,
    14-O-{[(1R, 2R, 4S)-4-Amino-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    14-O- {[(1R, 2R, 4R)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4S) diastereomer thereof,
    14-O-([(1R, 2R, 4S)-4-Aminomethyl-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    14-O-{[(1R, 2R, 4S)-4-[(2,2-Dimethyl-propylamino)-methyl]-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    14-O-{[(1R, 2R, 4S)-2-Hydroxy-4-[(2,2,2-trifluoro-acetylamino)-methyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 4R) diastereomer thereof,
    14-O-{[(1R, 2R, 3S)-2-Hydroxy 3-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer thereof,
    14-O-{[(1R, 2R, 3S)-3-(2-Amino-ethyl)-2-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 3R) diastereomer thereof,
    14-O-{[(1R, 2R, SR)-5-Hydroxy-2-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    14-O-{[(1R, 2R, 5R)-2-(2-Amino-ethyl)-5-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    14-O-{[(1R, 2R, 5R)-2-[2-(2,2-Dimethyl-propylamino)-ethyl]-5-hydroxy-cyclopentylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S) diastereomer thereof,
    14-O-{[(1R, 2R, 5R)-5-Amino-2-hydroxy-cycloheptylsulfanyl]-acetyl}-mutilin and the (1S, 2S, 5S), (1R, 2R, 5S)- (1S, 2S, 5R) diastereomers thereof,
    14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Amino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Formylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    14-O-{[(1R, 2S, 3S, 5R, 6S)-6-(2-Amino-acetylamino)-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    14-O-{[(1R, 2S, 3S, 5R, 6S)-6-Cyclopropylamino-2-hydroxy-bicyclo[3.2.0]hept-3-ylsulfanyl]-acetyl}-mutilin and the (1S, 2R, 3R, 5S, 6R) diastereomer thereof,
    14-O-{[4-Acetylamino-1-hydroxy-octahydropentalen-2-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof, and
    14-O-{[4-Acetylamino-2-hydroxy-octahydropentalen-1-ylsulfanyl]-acetyl}-mutilin and the diastereomers thereof.
  7. A compound according to any of claims 1 to 6 in the form of a salt and/or solvate.
  8. A compound according to any of claims 1 to 7 for use as a pharmaceutical drug substance.
  9. A method of treatment of diseases mediated by microbes which comprises administering to a subject in need of such treatment an effective amount of a compound of any one of claims 1 to 7.
  10. A pharmaceutical drug composition comprising a compound of any one of claims 1 to 7, in association with at least one pharmaceutical excipient.
  11. A pharmaceutical drug composition according to claim 10, further comprising another pharmaceutically active agent.
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